LUPUS ERITEMATOSO SISTEMICO

NEW ENG J MED.Volume 358:929-939
February 28, 2008
Number 9
Systemic Lupus Erythematosus
Anisur Rahman, Ph.D., and David A. Isenberg, M.D.

To the clinician, systemic lupus erythematosus is important because it is a potentially fatal disease that is easily confused with many other disorders. To the immunologist, lupus is intriguing because all the key components of the immune system are involved in the underlying mechanisms of the disease. This review describes these mechanisms and shows how knowledge of the pathogenesis of lupus facilitates its treatment.
The prevalence of lupus ranges from approximately 40 cases per 100,000 persons among Northern Europeans to more than 200 per 100,000 persons among blacks.1 In the United States, the number of patients with lupus exceeds 250,000. The life expectancy of such patients has improved from an approximate 4-year survival rate of 50% in the 1950s2 to a 15-year survival rate of 80% today.3 Even so, a patient in whom lupus is diagnosed at 20 years of age still has a 1 in 6 chance of dying by 35 years of age, most often from lupus or infection.4 Later, myocardial infarction and stroke become important causes of death.4 This bimodal pattern of mortality in lupus was recognized more than 30 years ago.5
The diverse presentations of lupus range from rash and arthritis through anemia and thrombocytopenia to serositis, nephritis, seizures, and psychosis. Lupus should be part of the differential diagnosis in virtually any patient presenting with one of these clinical problems, especially in female patients between 15 and 50 years of age.
Genetic and Epidemiologic Factors
Since 90% of patients with lupus are female, an important role for female hormones6 seems likely, but a protective role for male hormones or an effect of genes on the X chromosome is also possible. In a blinded, randomized, controlled trial, menopausal women with lupus who received hormone-replacement therapy containing conjugated estrogens and progesterone had a risk of a mild-to-moderate disease flare that was 1.34 times the risk among women who received placebo (P=0.01).7 However, trials of hormonal treatments for lupus, such as dehydroepiandrosterone, have been disappointing.8 It is unclear how sex hormones could promote lupus.
Many drugs cause a variant of lupus called drug-induced lupus. The best known of these drugs are procainamide, hydralazine, and quinidine. Patients with drug-induced lupus usually present with skin and joint manifestations; renal and neurologic features are very rare.9 An antecedent viral-like illness may occur at the onset of lupus or immediately before a flare. Identifying a particular causative virus has proved challenging. Epstein–Barr virus (EBV) may be important, since a temporal association between the onset of lupus and the occurrence of EBV infection has been reported. A case–control study involving children and young adults showed that anti-EBV antibodies were present in 99% and EBV DNA was present in 100% of patients with lupus — much higher proportions than those in the control group.10 Ultraviolet radiation is the most obvious environmental factor linked to lupus. A photosensitive rash is a criterion of the American College of Rheumatology for the classification of the disease.11,12
The concordance rate for lupus is 25% among monozygotic twins and approximately 2% among dizygotic twins13; these rates indicate that a genetic contribution is important, but it is not sufficient to cause the disease. Many genes that probably contribute to lupus have been identified by means of whole-genome scans from families in which multiple members have lupus.14,15 Eight susceptibility loci that have been identified in these studies . Genes of the major histocompatibility complex (MHC), particularly HLA-A1, B8, and DR3, have been linked to lupus.16 The response of a T lymphocyte to an antigen is triggered when a receptor molecule on the surface of the T cell recognizes a complex formed by the antigen and an MHC peptide on the surface of an antigen-presenting cell. Different types of cells within the immune system, such as B cells, macrophages, and dendritic cells, can function as antigen-presenting cells. The MHC genotype determines which MHC molecules are available to the antigens that are present and thus how well the antigens can be recognized by T cells. For this reason, particular MHC genes are associated with a risk of an immune response to self-antigens and hence a risk of diseases such as lupus.
Null alleles that cause a deficiency of one of the early complement components — C1q, C2, or C4 — are a strong risk factor for lupus.17 Family studies have identified genes that are more likely to occur in patients with lupus than in their healthy relatives.14 Many of these genes encode components of the immune system. For example, a Scandinavian study identified strong linkage between lupus and single-nucleotide polymorphisms in two interferon-related genes (those encoding tyrosine kinase 2 and interferon regulatory factor 5).18
Wakeland and colleagues14 have identified genetic loci that promote lupus in mice.19 These loci, designated Sle 1, Sle 2, and Sle 3, contain genes that mediate the loss of immunologic tolerance to nuclear autoantigens, B-cell hyperactivity, and T-cell dysregulation, respectively.14 The Sle 1 cluster contains genes similar to those in regions 1q21–23 and 1q41 of human chromosome 1 that have been linked to lupus in humans.14
Autoantibodies in Lupus
The affected organs in lupus that have been studied most intensively are the kidneys and the skin. In both cases, there is inflammation and the deposition of antibodies and complement. In 1967, kidneys from patients with lupus nephritis were shown to contain antibodies that bound native, double-stranded DNA.20 These antibodies are autoantibodies; that is, they bind a normal constituent — in this case, double-stranded DNA — of the patient's cells and tissues. The importance of anti–double-stranded DNA antibodies in the pathogenesis of lupus has been confirmed.21 Anti–double-stranded DNA antibodies are highly specific for lupus; they are present in 70% of patients with lupus but in less than 0.5% of healthy people or patients with other autoimmune diseases such as rheumatoid arthritis.22 Levels of anti–double-stranded DNA antibodies in serum tend to reflect disease activity,23 but not in all patients. Among patients who have both elevated levels of anti–double-stranded DNA autoantibodies and clinically quiescent disease, 80% have disease that becomes clinically active within 5 years after the detection of elevated levels of these antibodies.24
In a study of renal-biopsy specimens obtained from patients with lupus at autopsy,25 Mannik et al. detected IgG that bound to a number of non-DNA antigens, including Ro (a ribonucleoprotein complex), La (an RNA-binding protein), C1q (a subunit of the C1 complement component), and Sm (nuclear particles consisting of several different polypeptides). The detection of antibodies to these antigens in autopsy specimens does not prove that they play a role in the development of lupus nephritis. Rather than cause the inflammation, these autoantibodies may establish themselves in tissue only after the apoptosis of cells in inflamed kidney tissue exposes nuclear antigens. The strongest evidence concerning the mechanism of lupus nephritis relates to anti–double-stranded DNA, anti-nucleosome, and anti–-actinin antibodies (see below).
Although anti–double-stranded DNA antibodies are the most extensively studied autoantibodies in lupus, others play a role in clinical manifestations, particularly in autoimmune hemolytic anemia, thrombocytopenia, skin disease, and neonatal lupus. Table 2 lists common autoantibodies in lupus and the evidence that they are pathogenic; some are described in more detail below.
The presence of anti-Ro antibodies, anti-La antibodies, or both in pregnancy confers a 1 to 2% risk of fetal heart block. Ro antigens are exposed on the surface of fetal (but not maternal) cardiac myocytes as the heart undergoes remodeling by apoptosis, and maternal anti-Ro antibodies that cross the placenta interact with these antigens. The maternal autoantibodies damage the conducting tissues of the fetal heart.41,43 The absence of an effect on the mother's heart shows the importance of both the autoantibody and exposure of the antigen on cardiac tissue.
Antibodies against the N-methyl-D-aspartate (NMDA) receptor may be important in central nervous system lupus.27 NMDA is an excitatory amino acid released by neurons. Kawal and colleagues showed that in patients with lupus, the serum with antibodies against DNA and NMDA receptors caused cognitive impairment and hippocampal damage when given intravenously to mice. They also showed that anti–NMDA-receptor antibodies are present in the brain tissue of patients with cerebral lupus.27
Both anti-Ro and anti-nucleosome antibodies may play a role in cutaneous lupus. Anti-Ro antibodies are associated with an increased risk of the development of a photosensitive rash.42 Anti-nucleosome antibodies have been detected in skin-biopsy specimens obtained from a minority of patients with active renal lupus, and these patients had no rash.36
Autoantibody-mediated destruction of red cells and platelets is important in the hemolytic anemia and thrombocytopenia that can occur in patients with lupus.54 Pujol et al.55 detected antiplatelet antibodies in the serum of 56 of 90 patients with lupus. A total of 29 of 90 patients had thrombocytopenia, and in these patients there was a strong correlation between thrombocytopenia and the presence of antiplatelet antibodies.55
Tissue Damage by Autoantibodies in Lupus
Most studies of autoantibody-mediated tissue damage in patients with lupus have focused on the role of anti–double-stranded DNA antibodies in patients with lupus nephritis. There are two main theories; both stress that the binding of antibodies to double-stranded DNA itself is probably not the most critical determinant of tissue damage. Extracellular double-stranded DNA occurs mainly in the form of nucleosomes, which are fragments of chromatin that cells release when they undergo apoptosis. Berden and colleagues have proposed that pathogenic anti–double-stranded DNA autoantibodies in patients with lupus bind to nucleosomes that have entered the bloodstream; in turn, these antibody–nucleosome complexes settle in the renal glomerular basement membrane.56 These immune complexes activate complement, which initiates the glomerulonephritis. This series of events has been demonstrated in animal models.39,40 Furthermore, IgG antibodies have been shown, by means of electron microscopy, to colocalize with extracellular chromatin in lupus nephritis in humans and mice.37,38 Also relevant is the detection of anti-nucleosome antibodies in the blood and inflamed tissues of patients with lupus.26,36
The second model proposes that anti–double-stranded DNA, anti-nucleosome antibodies, or both cross-react with proteins in the kidney; thus, they have a direct pathogenic effect on renal cells. This is an example of polyreactivity, whereby the same antibody can bind to antigens with different structures because they have similar surface shapes (so-called shared epitopes) or areas of similar charge. Among possible target antigens in the kidney, attention is currently focused on -actinin. This protein is critical for maintaining the function of renal podocytes, which are constituents of the glomerular filtration barrier.57 Two studies have shown that mouse monoclonal anti-DNA antibodies that cross-reacted with -actinin (a protein that cross-links actin, a component of the cytoskeleton) were pathogenic, whereas monoclonal anti-DNA antibodies that did not cross-react with -actinin were nonpathogenic.52,53 Pathogenicity was judged according to whether the antibodies caused proteinuria and histologic changes of glomerulonephritis after passive transfer into recipient mice.52,58 Although anti–-actinin antibodies are not specific for lupus, these antibodies, when present in the serum of patients with lupus, can serve as a marker of renal involvement.28,51 The detection of anti--actinin antibodies has not been reported in specimens obtained from renal biopsies in patients with lupus.
The Role of T Cells
Autoantibodies can occur in healthy people without causing harm, and they may play a protective role.59 Pathogenic autoantibodies in patients with lupus have particular properties that enable them to cause disease. Clinical investigations and studies in laboratory mice have shown that IgG antibodies with high-affinity binding to double-stranded DNA tend to be more strongly associated with tissue damage than IgM or lower-affinity IgG antibodies.33,34,60 Production of these high-affinity IgG antibodies is "driven" by antigen. The term "antigen-driven" refers to a process in which antigen binds immunoglobulin on the surface of B lymphocytes, thereby stimulating the cells to proliferate. The higher the affinity of the surface immunoglobulin for the antigen, the more strongly the cells are stimulated and the more they proliferate. In the presence of the stimulating antigen, there is a continuous selective pressure favoring B cells that display on their surface and secrete immunoglobulins with high affinity for that antigen. In general, this antigen-driven process can occur only in B lymphocytes that are being stimulated by T lymphocytes as well as by antigen. This process is known as T-lymphocyte help.
The concept of T-lymphocyte help is critical in understanding the pathogenesis of lupus. Each T cell carries a surface-receptor molecule with the ability to interact best with one particular antigen when it is presented to the T-cell receptor in a complex with an MHC molecule on the surface of an antigen-presenting cell. Presentation of the antigen–MHC complex alone is not enough to stimulate the T cell. As shown in Figure 1, the antigen-presenting cell must also make a second molecular interaction with the T lymphocyte through costimulation. There are several different costimulatory molecular pairs, including the CD40–CD40 ligand and CD28–B7, which can generate the second signal required for T-cell activation. Agents that block costimulation can inhibit any immune response that depends on T-cell help. Since T-cell help is critical in lupus, both the anti-CD40 ligand61 and cytotoxic T-lymphocyte–associated protein 4 IgG1 (CTLA-4–Ig),62 a molecule that blocks the CD28–B7 interaction, are potential treatments for lupus. The prospects for these treatments are reviewed elsewhere.63
The antigen-presenting cell binds antigen in a complex with a molecule from the major histocompatibility complex (MHC) on its surface. This complex interacts with the T-cell receptor (TCR). The effect on the T cell depends on the interaction between other molecules on the surfaces of the two cells. Two alternative interactions are shown: B7 with CD28, which is stimulatory, and B7 with cytotoxic T-lymphocyte–associated protein 4 (CTLA-4), which is inhibitory. If the positive signal caused by the CD28–B7 interaction dominates, the T cell is activated, leading to cytokine release, B-cell help, and inflammation. If the negative signal caused by the CTLA-4–B7 interaction dominates, activation is suppressed.
Figure 2 shows a B cell and a T cell interacting and stimulating each other. T-cell cytokines affect B cells by stimulating cell division, switching antibody production from IgM to IgG,64 and promoting a change in the molecular sequence of the secreted antibody so that it binds more strongly to the driving antigen.65 Thus, T-cell help makes possible the production of high-affinity IgG autoantibodies. These kinds of antibodies are closely linked to tissue damage in lupus.33,34,60,66 The autoantigen-specific B cells and T cells that interact to produce injurious autoantibodies are absent in healthy people. Several mechanisms could account for the absence of such cells. These mechanisms include removal (deletion) of the autoreactive B cells, inactivation of the cells so that they remain in the body but are anergic, or a change in the light chain of the antibody expressed by an autoreactive B lymphocyte (so-called receptor editing) such that the antibody loses the ability to bind autoantigen. The use of certain light-chain genes by populations of B cells from patients with lupus indeed differs from the light-chain repertoire in healthy people; this difference could be due to aberrant receptor editing.67
The B cell acts as an antigen-presenting cell, with costimulation being obtained through the interaction between CD40 and the CD40 ligand. This interaction stimulates the T cell to produce a number of cytokines, some of which act on the B cell to promote antibody formation. MHC denotes major histocompatability complex, TCR T-cell receptor, and TNF tumor necrosis factor.
Histones constitute the protein core of a nucleosome, around which the DNA winds. Lu and colleagues68 showed that the histone-derived peptides H2B10-33, H416-39, H471-94, H391-105, H2A34-48, and H449-63 stimulated T cells from patients with lupus (but not from healthy people) to produce cytokines, and very similar peptides also stimulated T cells from lupus-prone mice. The authors suggested that stimulation of these peptide-specific helper T cells would allow them to help B cells that also respond to antigenic epitopes derived from nucleosomes. Thus, the interaction between these B lymphocytes and T lymphocytes could lead to the production of high-affinity pathogenic autoantibodies. Nucleosomes carry both T-cell and B-cell epitopes, and anti-nucleosome antibodies are present and play a pathogenic role in patients with lupus.26,39,40,56
Regulatory T cells in humans and mice suppress the activation of helper T cells and B cells. Some investigators have reported a reduction in the number or function — or both — of regulatory T cells in patients with lupus and in lupus-prone mice.69,70 Regulatory T cells from patients with active lupus have a reduced ability to suppress the proliferation of helper T cells, as compared with regulatory T cells from patients with inactive lupus or healthy controls.70 Kang et al. found that some of the immunogenic histone peptides they had previously identified promoted the development of regulatory T cells and delayed the development of nephritis in lupus-prone mice. The most potent effect was seen with peptide H471-94.71
Source of the Autoantigens in Lupus
The obvious source of nucleosomes is the cellular debris released as a result of apoptosis. During apoptosis, blebs of cellular material form on the surface of the dying cell. Antigens that are normally buried within the cells are exposed on the surface of these blebs (Figure 3), and they may trigger an immune response. These exposed antigens include nucleosomes, Ro 62, Ro 50, La, and anionic phospholipids.72 Antibodies to these antigens occur commonly in patients with lupus.
Apoptosis of keratinocytes exposed to ultraviolet light is illustrated. The different constituents of developing small and large surface blebs during apoptosis are shown. PARP denotes poly–ADP–ribose polymerase.
The removal of apoptotic debris is abnormal in patients with lupus.73 In vitro, phagocytes from patients with lupus were shown to engulf far less apoptotic material than phagocytes from healthy people during a 7-day culture period.74 C1q plays a role in phagocytosis by binding to cell debris, which can then be engulfed by macrophages that have surface C1q receptors. Thus, a deficiency of complement may be an important reason for the poor "waste disposal" seen in lupus. Homozygous deficiencies of C1q, C2, and C4 are rare disorders, but the presence of any of these genetic conditions is a strong predisposing factor for lupus.17 In C1q knockout mice, a lupuslike renal disease develops; kidney-biopsy specimens from mice with this condition reveal multiple apoptotic fragments.75 Davies and colleagues reported reduced clearance of immune complexes through the spleen in a patient with C2 deficiency and lupus; this was corrected by restoring the C2 levels with the use of transfusions of fresh-frozen plasma.76
Cytokines in Lupus
The role of tumor necrosis factor (TNF-) in lupus is controversial. This cytokine may be protective in patients with lupus, since giving TNF- to lupus-prone NZB/W F1 mice delayed the development of lupus.77 The protective effect is specific to that mouse strain, and the mechanism is unknown. In some patients with rheumatoid arthritis who were treated with anti–TNF- antibodies, anti–double-stranded DNA antibodies developed,78 and lupus developed in a few of these patients.79 One group has shown that the balance between TNF- and the soluble inhibitors (TNF-soluble receptor 75kDa and TNF-soluble receptor 55kDa) is altered in favor of the inhibitors in active lupus; this provides support for the idea that low TNF- activity is associated with increased disease activity in lupus.80 By contrast, the level of TNF- messenger RNA was high in kidney-biopsy specimens from patients with lupus nephritis.81 Aringer et al. reported that giving the anti–TNF- antibody agent infliximab to six patients with lupus led to resolution of joint swelling in three patients with arthritis and the reduction of urinary protein loss by 60% in four patients with renal lupus.82
Serum levels of interleukin-10 are consistently high in patients with lupus, and they correlate with the activity of the disease.83 Interleukin-10 has a number of biologic effects, including stimulation of polyclonal populations of B lymphocytes. Blocking this cytokine could reduce the production of pathogenic autoantibodies. In an open trial of 20 mg of a mouse anti–interleukin-10 antibody administered daily in six patients for 21 days, skin and joint symptoms improved in all the patients, and this improvement was maintained at the 6-month follow-up assessment.84
Serum levels of interferon- are also elevated in patients with active lupus,85 and microarray studies showed that 13 genes regulated by interferon were up-regulated in peripheral-blood mononuclear cells from patients with lupus, as compared with similar cells from healthy controls.86 In studies of lupus-prone NZB/W F1 mice, nephritis developed 15 to 20 weeks earlier in mice continuously exposed to interferon- from a young age than in control mice not subject to this exposure.87 Anti-interferon drugs may be the next anticytokine agents to be developed as treatments for patients with lupus.
The B-lymphocyte stimulator is a member of the TNF-ligand superfamily. It promotes the proliferation and survival of B lymphocytes. Circulating levels of B-lymphocyte stimulator are elevated in several other conditions, including rheumatoid arthritis and Sjögren's syndrome, as well as in lupus. The overexpression of B-lymphocyte stimulator has been detected in both humans with lupus and lupus-prone mice. Stohl et al. reported elevated levels of soluble B-lymphocyte stimulator in serum and on peripheral-blood mononuclear cells in up to 50% of patients with active lupus.88 Levels of B-lymphocyte stimulators correlated with levels of anti–double-stranded DNA antibodies in serum and decreased in nine patients who were treated with high-dose corticosteroids. Elevated levels of B-lymphocyte stimulators may thus be associated with the increased activity of lupus in some patients, and the use of anti–B-lymphocyte stimulator agents may be a useful therapeutic approach.
Implications for Treatment
Figure 4 summarizes the pathogenesis of lupus and the targets of some new drugs that are currently being evaluated in clinical trials. If autoantibodies are the proximate agents of tissue damage in patients with lupus, then treatments aimed at reducing autoantibody levels could be effective. Two trials31,32 have shown that a strategy of increasing doses of corticosteroids in response to a specified increase in levels of anti–double-stranded DNA antibodies leads to lower mean levels of such antibodies and reduced frequency of severe flares of disease, but one study indicated that the side effects of corticosteroids were a problem.31 Rituximab89 and abetimus sodium90 have been used as specific methods of reducing levels of anti–double-stranded DNA. Rituximab is nonspecific; that is, it is an antibody against CD20, which is found on the surface of all mature B cells. Abetimus sodium is designed to deplete only B lymphocytes that produce anti–double-stranded DNA antibodies because its four surface oligonucleotides can engage surface anti–double-stranded DNA antibodies on those cells, but it has no epitopes to allow binding of helper T cells. The B cells therefore undergo apoptosis rather than proliferation, but it is not clear whether this depleting mechanism occurs in patients. Abetimus sodium may also work by forming complexes with anti–double-stranded DNA antibodies, which are then cleared from the circulation.91
This simplified diagram, which is based on our increased understanding of the immunologic events thought to occur in lupus, indicates the targets of current therapeutic interventions. APC denotes antigen-presenting cell, BLyS B-lymphocyte stimulator, CTLA-4–Ig cytotoxic T-lymphocyte–associated protein 4 IgG1, and TACI-Ig transmembrane activator and CAML interactor immunoglobulin (CAML denotes calcium modulator and cyclophilin ligand).
Several case series suggest that rituximab is helpful in treating lupus.89,92 The use of a monoclonal anti-CD22 antibody (which also targets B cells)93 is being studied in a clinical trial, and the survival and proliferation of B cells can also be modulated with the use of anti–B-lymphocyte stimulator.88,94 A large trial showed that abetimus sodium was not superior to placebo in an analysis of the primary outcome measure (time to renal flare) for the whole study group, but in post hoc analyses, the drug was superior to placebo in a subgroup analysis of patients who had serum antibodies with high affinity for the drug.90
Anti-CD40 ligand61 and CTLA-4–Ig62 directly target the interaction between T cells and antigen-presenting cells by inhibiting costimulation. Peptides derived from pathogenic anti-DNA antibodies may be useful in generating anti-idiotypic responses to autoantibodies and thus suppressing their pathogenic effects.95 Trials of anti–TNF- antibody82 and anti–interleukin-10 antibody84 are described above.
Summary
Pathogenic autoantibodies are the primary cause of tissue damage in patients with lupus. The production of these antibodies arises by means of complex mechanisms involving every key facet of the immune system. Many different elements of the system are potential targets for therapeutic drugs in patients with lupus.
References
Johnson AE, Gordon C, Palmer RG, Bacon PA. The prevalence and incidence of systemic lupus erythematosus in Birmingham, England: relationship to ethnicity and country of birth. Arthritis Rheum 1995;38:551-558. [ISI][Medline]
Merrell M, Shulman LE. Determination of prognosis in chronic disease, illustrated by systemic lupus erythematosus. J Chronic Dis 1955;1:12-32. [CrossRef][Medline]
Abu-Shakra M, Urowitz MB, Gladman DD, Gough J. Mortality studies in systemic lupus erythematosus: results from a single center. II. Predictor variables for mortality. J Rheumatol 1995;22:1265-1270. [ISI][Medline]
Gladman DD, Urowitz MB. Prognosis, mortality and morbidity in systemic lupus erythematosus. In: Wallace DJ, Hahn BH, eds. Dubois' lupus erythematosus. 7th ed. Philadelphia: Lippincott Williams & Wilkins, 2007:1333-53.

DIABETES Y OBESIDAD INFANTIL

NEW ENG J MED.Volume 357:2325-2327
December 6, 2007
Number 23
Childhood Obesity — The Shape of Things to Come
David S. Ludwig, M.D., Ph.D.
Last week, I met with the G. family in the Optimal Weight for Life (OWL) clinic at my hospital. One of the parents was overweight, and the other was obese. The five children were more severely obese and had numerous weight-related complications — one had evidence of fatty liver, one had high blood pressure, two had gastroesophageal reflux, two had orthopedic problems, three had marked insulin resistance, four had dyslipidemia, and all had emotional problems related to their weight.
Sadly, this family might be a microcosm of 21st-century America: if we don't take steps to reverse course, the children of each successive generation seem destined to be fatter and sicker than their parents. How will obesity affect the physical and psychological well-being of children in coming decades? What effects will childhood obesity have on life expectancy, the national economy, and our society? To explore these questions, one might view the obesity epidemic as consisting of four overlapping phases.
Phase 1 began in the early 1970s and is ongoing: average weight is progressively increasing among children from all socioeconomic levels, racial and ethnic groups, and regions of the country. Today, about one in three children and adolescents is overweight (with a body-mass index, or BMI, in the 85th to 95th percentile for age and sex) or obese (BMI above the 95th percentile), and the proportion approaches one in two in certain minority groups.1 Though it has attracted much attention from the medical profession and the public, childhood obesity during this phase has actually had little effect on public health, because an obese child may remain relatively healthy for years.
Phase 2, which we are now entering, is characterized by the emergence of serious weight-related problems.2 The incidence of type 2 diabetes among adolescents, though still not high, has increased by a factor of more than 10 in the past two decades and may now exceed that of type 1 diabetes among black and Hispanic adolescents. Fatty liver associated with excessive weight, unrecognized in the pediatric literature before 1980, today occurs in about one in three obese children. Other obesity-related complications affecting virtually every organ — ranging from crippling orthopedic problems to sleep apnea — are being diagnosed with increasing frequency in children (see table). There is also a heavy psychosocial toll: obese children tend to be socially isolated and have high rates of disordered eating, anxiety, and depression. When they reach adulthood, they are less likely than their thinner counterparts to complete college and are more likely to live in poverty.
Complications of Childhood Obesity.
It may take many years to reach phase 3 of the epidemic, in which the medical complications of obesity lead to life-threatening disease. As Baker et al. (pages 2329–2337) and Bibbins-Domingo et al. (pages 2371–2379) report in this issue of the Journal, overweight or obesity in childhood or adolescence increases the risk of coronary heart disease (CHD) in adulthood; by 2035, Bibbins-Domingo et al. predict, the prevalence of CHD will have increased by 5 to 16%, with more than 100,000 excess cases attributable to increased obesity among today's adolescents. Preliminary data from Canada suggest that adolescents with type 2 diabetes will be at high risk for limb amputation, kidney failure requiring dialysis, and premature death. In some, fatty liver will progress to hepatitis and cirrhosis, which may remain asymptomatic until irreversible organ damage has occurred. Poverty and social isolation would complicate the timely identification and management of such problems. Shockingly, the risk of dying by middle age is already two to three times as high among obese adolescent girls as it is among those of normal weight, even after other lifestyle factors are taken into account.3 My colleagues and I have predicted that pediatric obesity may shorten life expectancy in the United States by 2 to 5 years by midcentury — an effect equal to that of all cancers combined.4
Without effective intervention, phase 4 of the epidemic will entail an acceleration of the obesity rate through transgenerational mechanisms. Obese children tend to be heavy in adulthood, in part because obesity-promoting habits persist. In addition, carrying excessive weight early in life may elicit irreversible biologic changes in hormonal pathways, fat cells, and the brain that increase hunger and adversely affect metabolism. Furthermore, adult obesity and its complications appear to increase the risk of obesity and its complications in offspring through nongenetic influences, a phenomenon termed perinatal programming. For example, a recent study found that maternal hyperglycemia during pregnancy strongly predicted BMI in offspring at 5 to 7 years of age, after adjustment for maternal weight gain and birth weight.5
Currently, the economic costs of pediatric obesity in the United States are relatively small — probably several hundred million dollars annually. Without effective intervention, the costs of obesity might well become catastrophic, arising not only from escalating medical expenses but also from diminished worker productivity, caused by physical and psychological disabilities. Future economic losses could mean the difference between solvency and bankruptcy for Medicare, between expanding and shrinking health care coverage, and between investment in and neglect of our social infrastructure, with profound implications for our international competitiveness. The human costs would be incalculable.
Like global warming, the obesity epidemic is a looming crisis that requires action before all the scientific evidence is in. And as with climate change, some have questioned experts' forecasts, doubting the far-reaching impact of obesity, though skepticism is gradually being overcome by accumulating data. Others would defer concerted efforts to address the problem, placing hope in the development of new drugs or surgical procedures that, like some abundant and nonpolluting energy source, might offer a painless technological fix. Or they argue that the costs of action are too great, not recognizing that our survival depends on solving the problem. But I believe that obesity differs in one important respect from global warming: simple solutions are available, and with a comprehensive national strategy, we may be able to implement them without great sacrifice.
Certainly, we have much to learn about the regulation of body weight. Low-fat diets have yielded disappointing results, and very-low-carbohydrate diets appear to be more effective only in the short term. Novel approaches that focus on the quality rather than the ratio of macronutrients appear promising, and other areas warrant study, including the effects of sleep deprivation, stress, infectious agents, and endocrine-disrupting environmental toxins on weight. Unfortunately, the U.S. government has thus far invested only a fraction of a cent in research for every dollar that obesity costs society. And although broad consensus exists regarding the dietary and lifestyle habits needed to prevent and treat childhood obesity, we lack anything resembling a comprehensive strategy for encouraging children to eat a healthful diet and engage in physical activity. Such a strategy would include legislation that regulates junk-food advertising, provides adequate funding for decent lunches and regular physical activities at school, restructures the farm-subsidies program to favor nutrient-dense rather than calorie-dense produce, and mandates insurance coverage for preventing and treating pediatric obesity.
Parents must take responsibility for their children's welfare by providing high-quality food, limiting television viewing, and modeling a healthful lifestyle. But why should Mr. and Ms. G.'s efforts to protect their children from life-threatening illness be undermined by massive marketing campaigns from the manufacturers of junk food? Why are their children subjected to the temptation of such food in the school cafeteria and vending machines? Why don't they have the opportunity to exercise their bodies during the school day? And why must Mr. and Ms. G. fight with their insurance company for reimbursement to cover the costs of their children's care at the OWL clinic? Fortunately, with the exercise of both personal and social responsibility, we have the power to choose the shape of things to come.
References
Ogden CL, Carroll MD, Curtin LR, McDowell MA, Tabak CJ, Flegal KM. Prevalence of overweight and obesity in the United States, 1999-2004. JAMA 2006;295:1549-1555. [Free Full Text]
Ebbeling CB, Pawlak DB, Ludwig DS. Childhood obesity: public health crisis, common sense cure. Lancet 2002;360:473-482. [CrossRef][ISI][Medline]
van Dam RM, Willett WC, Manson JE, Hu FB. The relationship between overweight in adolescence and premature death in women. Ann Intern Med 2006;145:91-97. [Free Full Text]
Olshansky SJ, Passaro DJ, Hershow RC, et al. A potential decline in life expectancy in the United States in the 21st century. N Engl J Med 2005;352:1138-1145. [Free Full Text]

MAPA GENETICO DEL ORIGEN HUMANO

Confirman la teoría Out of Africa: la especie humana apareció en el sur de ese continente
Mapa genético detalla origen del hombre y la colonización del planeta
Científicos estudiaron el genoma de 51 grupos para detectar cambios en los descendientes
Descubren que el árbol genealógico europeo se divide en ocho ramas
Los expertos detectaron pequeñas diferencias en la secuencia de 3 mil 200 millones de pares base que son similares en los humanos
Washington, 21 de febrero. Un estudio genético para determinar el origen del hombre y la colonización de la Tierra sigue el camino del hombre desde África a todo el mundo de manera muy detallada, como no se había logrado anteriormente.
Jun Li y colegas de la Escuela Universitaria de Medicina de Stanford, en California, presentaron su investigación en la revista estadunidense Science (volumen 319, página 1,100), en su edición de mañana viernes, un día después de la publicación en la revista británica Nature de otros dos estudios genéticos, que confirman que el hombre pobló el mundo desde África.
El hombre partió de África hacia Medio Oriente y luego conquistó Europa, dice el grupo de Jun Li, con base en el análisis comparativo del genoma de 938 personas no emparentadas de 51 grupos que viven en todo el mundo.
Aún en la actualidad se pueden detectar sutiles variantes en el genoma de, por ejemplo, los vascos y franceses. También se diferencian los hombres de la Toscana de los del resto de Italia.
El estudio confirma una vez más que el mundo se fue poblando gradualmente desde el sur de África, lo que se conoce como hipótesis Out of Africa.
Nuevas ramas
La secuencia completa fue: sur de África, norte de África, Medio Oriente, Europa, centro y sur de Asia, Oceanía, América. Tras conquistar una región, un pequeño grupo de hombres partía en busca de nuevos horizontes, indicaron los expertos.
Jun Li y colegas incorporaron los datos de los 51 grupos en un nuevo árbol genealógico, que en la región europea se divide en ocho ramas: personas de Toscana, Cerdeña, Italia, País Vasco, franceses, escoceses de las islas Orkneyi, rusos y los pobladores de la región rusa de Adygei.
Sin embargo, indicaron que seguramente debe de haber en Europa muchos más grupos. Además, con los métodos disponibles en la actualidad no se pueden detectar grandes variantes entre muchos grupos de población europeos y sus antecesores.
La investigación también muestra que los habitantes de los pueblos San del sur de África están en la raíz del árbol genealógico. Posiblemente se podrían diferenciar aquí otros grupos, pero los investigadores dirigidos por Jun Li sólo tenían a disposición datos sobre representantes de 51 grupos en todo el mundo.
Para su estudio, los expertos aprovecharon pequeñas diferencias en el genoma, cuya secuencia de unos 3 mil 200 millones de pares de bases es similar en todos los hombres. Sin embargo, en algunos casos aparecen errores: entonces hay un cambio de un par de bases por otro.
Esta mutación, conocida como polimorfismo nucleótido simple (SNP, por sus siglas en inglés), en general no tiene consecuencias en el fenotipo, pero se transmite a la descendencia. A lo largo de mucho tiempo, en determinadas regiones aparece un distribución característica de estos SNP en el genoma.
Si un grupo emigra a otro continente, se lleva esta característica y la transmite a la escendencia.
Por lo tanto, su análisis revela muchos datos sobre el origen y los antecesores de un hombre. Los asiáticos tienen una distribución diferente de los SNP que los europeos o africanos.
Las diferencias aumentan por la distancia tanto temporal como espacial de los hombres primitivos del sur de África.
Jun Li y colegas utilizaron para su investigación material genético del Proyecto de Diversidad del Genoma Humano. LA JORNADA. 22 DE FEBRERO 2008

SINDROME DE TURNER

NEW ENG J MED.Volume 351:1227-1238
September 16, 2004
Number 12
Turner's Syndrome
Virginia P. Sybert, M.D., and Elizabeth McCauley, Ph.D.

Turner's syndrome, a disorder in females characterized by the absence of all or part of a normal second sex chromosome, leads to a constellation of physical findings that often includes congenital lymphedema, short stature, and gonadal dysgenesis.1,2,3 Turner's syndrome occurs in 1 in 2500 to 1 in 3000 live-born girls. Approximately half have monosomy X (45,X), and 5 to 10 percent have a duplication (isochromosome) of the long arm of one X (46,X,i(Xq)). Most of the rest have mosaicism for 45,X, with one or more additional cell lineages .
In the past decade, more has been learned about the natural history of Turner's syndrome, and recent molecular studies have identified some genes that may be involved in the clinical expression of the condition. This review summarizes current knowledge and makes recommendations for care on the basis of the medical literature and on our own experience with 532 live-born children and adults with karyotypically confirmed Turner's syndrome 4
Diagnosis
When Turner's syndrome is diagnosed prenatally, the diagnosis is usually based on the finding of fetal edema on ultrasonography; abnormal levels of human chorionic gonadotropin, unconjugated estriol, and alpha-fetoprotein on screening of maternal serum (triple screening); or abnormal results of fetal karyotyping performed because of advanced maternal age. Affected fetuses often abort spontaneously. A 45,X fetus identified prenatally and surviving to birth has a prognosis similar to that of a child in whom Turner's syndrome is diagnosed postnatally. In contrast, approximately 90 percent of fetuses in whom 45,X/46,XX or 45,X/46,XY mosaicism is diagnosed incidentally during the course of screening for advanced maternal age or maternal triple screening will likely have a normal phenotype, female or male, respectively, at birth. The risk of eventual gonadal failure in these children with mosaicism is unknown.5,6 In contrast, a child in whom 45,X/46,XX or 45,X/46,XY mosaicism is diagnosed after birth is usually identified because of phenotypic features suggestive of Turner's syndrome; such children have a prognosis similar to that for 45,X children.5
One fifth to one third of affected girls receive a diagnosis as newborns because of puffy hands and feet or redundant nuchal skin , the residual effect of cystic hygromas in utero. Turner's syndrome should be suspected in any newborn girl with edema or hypoplastic left heart or coarctation of the aorta, since the frequency of both conditions is increased among children with Turner's syndrome. Approximately one third of girls with Turner's syndrome receive the diagnosis in midchildhood on investigation of short stature. With the exception of familial short stature or constitutional delay, Turner's syndrome is the most common cause of short stature in otherwise healthy girls. In most other patients with Turner's syndrome, the condition is diagnosed either in adolescence when they fail to enter puberty or in adulthood because of recurrent pregnancy loss. The diagnosis should be excluded in any teenage girl with primary or secondary amenorrhea, especially if she is short.
Karyotyping of a blood sample is definitive in most cases. Detection of mosaicism depends on the proportion of cells present from the additional cell lineages. In routine karyotyping, 20 cells are counted, since this number is sufficient to detect mosaicism at a level of about 5 percent.
Mosaicism for a second, normal 46,XX cell population occurs in approximately 15 percent of girls with Turner's syndrome. Extensive searching for 46,XX cells in a girl with a 45,X karyotype is not necessary, since the detection of a normal cell lineage in fewer than 5 percent of cells does not change the prognosis or the management. Conversely, if the diagnosis of Turner's syndrome is suspected clinically but the result of routine testing is normal, increasing the number of cells counted to 100 and performing a skin biopsy for karyotyping of fibroblasts are indicated to rule out mosaicism for an abnormal cell lineage.
Girls with mosaicism for a cell population with a Y chromosome are at increased risk for gonadoblastoma (risk, 7 to 30 percent) in their streak gonads.7 Although the use of flow cytometry or DNA hybridization to search for Y-chromosome material has been suggested for all girls with a 45,X karyotype,8 clinical evidence indicates that such an approach is merited only in those with masculinization or mosaicism for an unidentified marker. The use of polymerase-chain-reaction testing for Y-chromosome sequences has a high false positive rate.9
Which chromosomal regions and genes account for the physical characteristics of Turner's syndrome remains uncertain 10,11,12 It has been hypothesized that the physical manifestations of Turner's syndrome are due either to the absence of two normal sex chromosomes before X-chromosome inactivation or to haploinsufficiency of genes in the pseudoautosomal regions of the X or Y chromosome, as well as to aneuploidy itself.13,14 Both the short arm and the long arm of the X chromosome contain genes important for ovarian function, and aneuploidy alone may lead to a reduction in the number and survival of oocytes.
POF2 denotes the premature ovarian failure gene, an unidentified gene hypothesized to be responsible for ovarian failure on the basis of the study of translocations. The POF1 gene is homologous to the diaphanous gene (DIAPH2) in the fruit fly. SHOX is located within PAR1.
Loss of interstitial or terminal long-arm material of the X chromosome (Xq) can result in short stature and primary or secondary ovarian failure.15 Deletions distal to Xq21 appear to have no effect on stature. In general, loss of the short arm (Xp) results in the full phenotype. Very distal Xp deletions are compatible with, but do not ensure, normal ovarian function.11,12 Loss of this region usually confers short stature and the typical skeletal changes, in part as a result of haploinsufficiency of the short stature–homeobox (SHOX) gene, located in the pseudoautosomal region of Y and Xp.16 The SHOX gene is probably not the only gene responsible for the skeletal features. Aneuploidy itself may contribute to growth failure.14 Loss of a region at Xp22.3 appears to be associated with the neurocognitive problems in Turner's syndrome.17 Loss of the testis-determining factor (SRY) gene locus on the short arm of the Y chromosome (e.g., 46,X,del(Yp)) also leads to the phenotype of Turner's syndrome, even without a 45,X cell population. A region on Xp11.4 has been proposed as critical for the development of lymphedema.18
There are some correlations between karyotype and phenotype (Table 1). Infants with a 45,X karyotype are the most likely to have congenital lymphedema. Patients with a karyotype of 45,X/46,XX or 45,X/47,XXX are the most likely to have spontaneous menarche and fertility.4,19 As a group, women with mosaicism for 45,X/46,XX are marginally taller than other women with Turner's syndrome. The presence of an isochromosome Xq suggests an increased risk for hypothyroidism and inflammatory bowel disease.3,4,20 The presence of a ring or marker chromosome confers an increased risk of mental retardation and atypical phenotypic features. Nonetheless, phenotypic predictions for a given patient that are based on karyotype are unreliable in patients with Turner's syndrome. Women with a 45,X karyotype have conceived; women with a 45,X/46,XX karyotype and a preponderance of 46,XX cells may have all the findings of the disorder.
Management
Growth
The mean birth length of infants with Turner's syndrome falls within the low end of the normal range. A decrease in growth velocity occurs as early as 18 months of age.21 Many patients will not be the shortest child in kindergarten but will have had a significant decrease in linear growth rate by third or fourth grade. Some present only when the normal pubertal growth spurt fails to occur. It is easy to misinterpret the absence of puberty and small size of these patients as due to constitutional delay; 104 of 150 patients who came to our attention as teenagers had had evidence of growth failure earlier in childhood that had been overlooked.4
A study of the efficacy of recombinant human growth hormone in patients with Turner's syndrome was initiated in 1983 in the United States22 and led to approval of this agent by the Food and Drug Administration in 1997. Treatment with recombinant human growth hormone is now standard in many centers, though physiologically significant alterations in growth hormone secretion have not been identified in patients with Turner's syndrome. Studies that followed treated patients to their final height23,24,25,26,27,28,29,30,31,32,33,34,35,36 have based therapeutic success on one of three measures: the mean final height of the treated group, as compared with a historical mean height of 143.2 cm37; the height achieved by each subject, as compared with her projected height on the basis of her centile on the Lyon curve (a growth chart specific to patients with Turner's syndrome)37 at the onset of treatment with recombinant human growth hormone; and the subject's predicted height, which was derived from midparental height.38 Only one published, nonrandomized study has included a concurrent control group.30 Two studies that include randomized control groups have been initiated — one in Canada and one at the National Institutes of Health. Only interim results in abstract form are available for the former39; the latter is ongoing.
Comparisons of the final heights of girls treated with recombinant human growth hormone with projected or predicted heights range from no gain to an increase of as much as 11.9 cm.22,23,24,25,26,27,28,29,30,31,32,33,34,35,36 Differences in ages at the commencement of treatment and differences in the doses and duration of therapy complicate analysis. The use of historical controls whose measurements led to the Lyon growth curves may not be valid for contemporary populations. For example, the mean adult height in 149 of our untreated patients is currently 148 cm, 4.8 cm taller than the mean adult height of the Lyon curve. Although one study suggested that all treated girls reached or exceeded their predicted adult height,22 other studies have not.33,38,39,40,41 The ideal dosing regimens and duration of treatment have not been established.36,42 It has been estimated that the cost of recombinant human growth hormone per centimeter of final gain in height is approximately $29,000.42
The short-term safety of treatment with recombinant human growth hormone in patients with Turner's syndrome appears to be acceptable. Increased insulin resistance and increased blood pressure have occurred during therapy and resolve on its cessation.43 The long-term effects of recombinant human growth hormone treatment on cardiovascular status,44 especially on aortic-root diameter, and the lifetime risk of type 2 diabetes are unknown. No systematic studies have examined whether treatment with recombinant human growth hormone improves the psychosocial outcomes and the quality of life of patients with Turner's syndrome.
Our view is that recombinant human growth hormone should be considered for every girl with Turner's syndrome. Parents and children should be told of the limitations of current knowledge about treatment and be given realistic expectations with respect to the resulting gain in height, so that they can make informed decisions.28 Most adults with Turner's syndrome cope successfully with their small stature.45
Weight management is an issue in patients with Turner's syndrome. Obesity is neither inherent nor unavoidable.46 Affected girls should be encouraged to engage in physical activities such as swimming, walking, and bicycling beginning in childhood and continuing throughout their lives.
Developmental and Behavioral Concerns
Most people with Turner's syndrome have normal intelligence. Approximately 10 percent of patients (Table 1), irrespective of karyotype, will have substantial developmental delays, need special education, and require ongoing assistance in adult life. The risk of mental retardation is highest among patients with a marker chromosome (66 percent) or a ring (X) chromosome (30 percent).4
Approximately 70 percent of patients with Turner's syndrome have learning disabilities affecting nonverbal perceptual motor and visuospatial skills.47,48,49 These deficits appear to be more common among patients with a 45,X karyotype than among those with a 45,X/46,XX karyotype.50 Better verbal and executive skills may be associated with inheritance of a paternally derived X chromosome,51 although these findings have not been corroborated. Findings on magnetic resonance imaging and positron-emission tomography have suggested the presence of nonspecific differences between patients with Turner's syndrome and controls, particularly in the right posterior regions of the brain.52,53 How these differences may relate to neurocognitive findings is unknown.
A meta-analysis of 13 studies involving 226 patients with Turner's syndrome and 142 controls identified deficits in visuospatial organization, social cognition, nonverbal problem-solving, and psychomotor functioning in the patients.49 Deficits in nonverbal memory,54 executive function,55,56 and attentional abilities47 are common. As with nonverbal learning disabilities,57 these deficits translate into problems with diverse activities such as mathematics,58,59 driving, multitasking, and social functioning.49,56 Spatial and math deficits appear early; problems with reading comprehension emerge as more complex academic demands are made. Attention-deficit–hyperactivity disorder is relatively common.60 Early cognitive testing and appropriate accommodations, such as tutoring; enrollment in small, structured classes; and the use of untimed testing, may be indicated.
Girls with Turner's syndrome have typical female-sex identification. Most affected women report being heterosexual, although they are less likely than their peers to have sexual relationships and do so at an older age.45,61 Prevalence rates of coexisting psychiatric diagnoses range from 2 to 10 percent,4,62 which are actually lower than the rate of 14 percent among the general population.63
Younger patients may have impaired peer relationships and anxiety and may be preoccupied with keeping things in order and inflexible regarding changes in their routine.64,65,66 They have relatively poor self-esteem, particularly in the social arena, as compared with both girls with short stature from other causes and girls with normal height.60,66,67
During adolescence, immaturity, social isolation, and anxiety are common.60,64 People with Turner's syndrome may misread social cues, facial expressions, and body language,47,65,68 contributing to awkwardness in social interactions; special training in recognizing social cues may be helpful.
Successful transition of these patients into the working world requires age-appropriate, not size-appropriate, expectations. During driver's training, many adolescents will require attention to be paid to their impaired navigational planning, visuomotor integration, and spatial and directional abilities. Most adults with Turner's syndrome report satisfaction with their lifestyle45,69; they have fewer social contacts than their peers but do not perceive themselves to be isolated.45 They react with appropriate depression and feelings of loss related to their physical limitations and usually cope well; their sense of self appears to be directly related to their health status.45
Women with Turner's syndrome are often employed at occupational levels below that predicted on the basis of their education and training. They may fail at jobs requiring a rapid response and multitasking, reflecting the effect of nonverbal learning disabilities.70 Nonetheless, many have successful professional careers.
Cardiovascular Concerns
The prevalence of congenital heart disease among patients with Turner's syndrome ranges from 17 to 45 percent, with no clear phenotype–genotype correlations.3,71,72 Death from cardiac causes is a serious concern.4,73 Coarctation of the aorta and bicuspid aortic valve are the most common structural malformations, followed by other left-sided defects. Hypertension, mitral-valve prolapse, and conduction defects also occur. Hypertension in the absence of structural cardiac malformations is usually not associated with arteriosclerotic heart disease or renal disease.4 The risk of hyperlipidemia and coronary artery disease in patients with Turner's syndrome is unclear.
Echocardiography is a mandatory part of the diagnostic workup for Turner's syndrome, since a physical examination may be inadequate to detect a bicuspid aortic valve.71 Use of magnetic resonance imaging as a screening tool for Turner's syndrome has not been standardized.
There have been more than 80 reports of aortic dissection in patients with Turner's syndrome.71,74,75,76 Coarctation of the aorta (unrepaired or repaired), bicuspid aortic valve, hypertension, or a combination of these findings, which are risk factors for aortic dissection, were present in 93 percent of these patients. The normal range of aortic-root diameters has not been established in patients with Turner's syndrome. The need for and frequency of repeated echocardiography for the assessment of the aortic-root diameter in those without structural cardiac abnormalities is unknown and should be individualized (Table 2).71,76
Endocrine Concerns
Hypothyroidism occurs in 15 to 30 percent of women with Turner's syndrome.4,20,77 The mean age at onset is in the third decade, though 5 to 10 percent of cases occur before adolescence. Acute thyroiditis is uncommon. Screening of thyroid function, including measurement of thyrotropin levels, should begin at about 10 years of age in asymptomatic patients. We do not monitor antithyroid antibody status, since the presence of these antibodies does not alter management.
Gonadal dysgenesis is a cardinal feature of Turner's syndrome; 90 percent of patients will require hormone-replacement therapy to initiate puberty and complete growth. In utero, the ovaries have a decreased number of primordial follicles; these appear to undergo premature apoptosis78 and are usually absent by adult life. The uterus may be small owing to a lack of estrogen; structural uterine abnormalities are rare. Dyspareunia sometimes occurs because of a small vagina or an atrophic vaginal lining.
The presence of normal gonadotropin levels in the first three to six months of life suggests that residual ovarian function exists but does not ensure that the initiation and progression of puberty will be normal. Gonadotropins are suppressed in childhood, even in those with gonadal dysgenesis. In many girls with Turner's syndrome, pubic and axillary hair will develop spontaneously, but changes of adrenarche are not indicative of ovarian function. Some girls have enough residual ovarian function for breast budding or vaginal spotting to occur, but secondary amenorrhea will develop. A minority will maintain ovulatory cycles for a time. Two fifths of girls with 45,X/46,XX mosaicism will have spontaneous menarche; however, ovarian failure usually ensues.4 If the status of ovarian function in adolescence is unclear, measurement of follicle-stimulating hormone, luteinizing hormone, and estradiol levels can help determine the need for hormone-replacement therapy.
Hormone-replacement therapy should be initiated at about the age of 14 years.4,79 Earlier treatment may result in a decrement in final height. Psychosocial issues and the patient's maturity and wishes also need to be considered. Girls who have received recombinant human growth hormone and who have completed most of their growth, as judged on the basis of bone age or growth velocity, may start hormone-replacement therapy at the age of 12 years if they wish.
There is no single formula for the use of hormone-replacement therapy.80 Several strategies are outlined in Table 3. After the first year, the use of a cycling regimen with a progestational agent is mandatory to minimize the risk of endometrial hyperplasia and uterine adenocarcinoma.
The effects of hormone-replacement-therapy on liver function, on bone density, and on the risk of hypertension, cancer, and obesity in patients with Turner's syndrome are uncertain. Although there have been very few reports of frank liver disease in women with Turner's syndrome, elevated liver enzymes have been reported,81 and the use of different forms of estrogen replacement may ameliorate or exacerbate this problem.82 There are currently insufficient data to make specific recommendations. There has not been an increased occurrence of breast cancer among patients with Turner's syndrome.83
Spontaneous fertility is rare among patients with Turner's syndrome and is most likely in women with mosaicism for a normal 46,XX cell lineage, a 47,XXX cell lineage, or very distal Xp deletions.19 These women have an increased risk of spontaneous pregnancy loss, twins, and aneuploidy in fetuses that are carried to term.4,19,84 Efforts to cryopreserve ovarian tissue are fairly new, and the applicability of such techniques to preserve fertility in women with Turner's syndrome may be compromised by a high rate of aneuploid gametes.
Physicians should discuss infertility issues and reproductive options with their patients and reassure them about their sexual function. It is important to acknowledge the sense of loss associated with infertility, on the part of both the patient and her parents. Pregnancy, by means of gamete intrafallopian transfer with donor eggs, has been attempted in women with Turner's syndrome, with a success rate equal to that in other infertile women. However, there have been five case reports of aortic dissection in women with Turner's syndrome who have undergone gamete intrafallopian transfer. Two of these cases may represent duplicate reports; inadequate details were provided to be certain. In a collected series,85 101 of 146 women with Turner's syndrome in whom gamete intrafallopian transfer was attempted became pregnant; none had aortic dissection. One woman had an aortic dissection before the procedure. Among 93 reports of women with Turner's syndrome who have become pregnant spontaneously, there have been no occurrences of aortic dissection.4
The prevalence of insulin resistance and type 2 diabetes may be increased in patients with Turner's syndrome. Among 257 patients in several large series, 18 (7 percent) had diabetes requiring treatment.4 Diabetes has developed in 11 of our 372 patients older than five years of age for whom we have information (type 1 in 3 and type 2 in 8).4 The majority of patients with Turner's syndrome and diabetes have adult-onset diabetes, and most are overweight. There is conflicting evidence regarding the effect of hormone-replacement therapy on glucose homeostasis in patients with Turner's syndrome and none regarding the long-term effects of recombinant human growth hormone.
Ophthalmologic and Otologic Concerns
Clinically significant strabismus occurred in 18 percent of our patients with Turner's syndrome, and ptosis in 13 percent.4 Cataracts and nystagmus also occur more commonly in patients with Turner's syndrome. Red–green colorblindness is found with the same frequency as in normal males.4,86 There should be a low threshold for referral to ophthalmologists for these patients.
The majority of infants and children with Turner's syndrome have recurrent otitis media, which is probably due to a combination of small, dysfunctional eustachian tubes and palatal dysfunction. This can be a major problem in early childhood, causing substantial complications and many sleepless nights. The frequency of ear infections decreases with age and growth of facial structures. Palatal dysfunction in these patients may be exacerbated by the removal of adenoids. Such surgery should be undertaken only after a careful evaluation of the patient's speech and palatal configuration.
Progressive sensorineural hearing loss is a major feature of Turner's syndrome in adults. Ninety percent of 44 adults with Turner's syndrome had sensorineural hearing loss. The loss was clinically significant in two thirds, and 27 percent required hearing aids.87 Five percent of children and 17 percent of adults in our clinic require hearing aids. The biologic basis is not known.
Gastrointestinal Manifestations
Feeding problems, gastroesophageal reflux, and failure to thrive occur in both breast-fed and bottle-fed infants with Turner's syndrome, possibly as a result of anatomical differences in the oropharynx as well as oral motor immaturity.88 There have been rare reports of a variety of symptomatic vascular malformations of the gastrointestinal tract. More common are instances of inflammatory bowel disease. In a series of 135 adults with Turner's syndrome, 2 had Crohn's disease, 2 had ulcerative colitis, and 2 had chronic diarrhea of unknown cause.89 More than half of patients with Turner's syndrome and inflammatory bowel disease who have been described in the literature have had an i(Xq) cell lineage. There may be an increased incidence of celiac disease among patients with Turner's syndrome; preliminary screening studies have shown elevated levels of IgA–antiendomysium and IgA–antigliadin antibodies in 2 to 10 percent of patients who were screened but symptomatic disease in only a few.3,90,91 The incidence of gallbladder disease may be higher than expected and is not associated with diabetes or obesity.4
Renal Manifestations
Structural renal malformations, including horseshoe kidney and duplication of the collecting system, are found in up to 40 percent of patients with Turner's syndrome.4,92 Whereas most structural malformations do not cause renal dysfunction, silent hydronephrosis resulting from obstruction of a duplicated collecting system may occur (present in 10 percent of our patients). Screening renal ultrasonography is necessary for all patients with Turner's syndrome.
Musculoskeletal Characteristics
Turner's syndrome is characterized by skeletal dysplasia, with short stature, mild epiphyseal dysplasia, and typical bony alterations. Dislocation of the patellae and chronic knee pain are common. Malformation of the ulnar head causes the typical increased carrying angle of the arm and may cause limited range of motion. Chondrodysplasia of the distal radial epiphysis (Madelung's deformity), typical of the Leri–Weill syndrome — the skeletal dysplasia associated with SHOX haploinsufficiency — is a rare complication. Congenital dislocation of the hip is common (occurring in 5 percent of patients), as is clinically significant scoliosis (occurring in 10 percent).4
It is unclear whether patients with Turner's syndrome have an increased risk of osteoporosis or fractures.93,94,95,96 Their bones appear osteopenic on radiographic evaluation, and their regional bone mass is often, but not always, below that of age-matched, but not height-matched, controls.95 No longitudinal studies have been done to establish whether the reduced bone mass is a nonprogressive feature of a general skeletal dysplasia or is analogous to the accelerated bone loss seen in postmenopausal women primarily as a result of estrogen deficiency. Both hormone-replacement therapy and recombinant human growth hormone treatment may improve regional bone mass.95,97 However, one study found no differences among patients treated with growth hormone, estrogen replacement, or both and an age-matched group of untreated patients with Turner's syndrome.97
Dermatologic Concerns
It may take several years for the congenital puffiness of the hands and feet to resolve in patients with Turner's syndrome. In rare cases, pedal edema persists or recurs in late childhood, at the time of ovarian hormone-replacement treatment, or later. There is an increased number of typical melanocytic nevi that are not clinically or histologically unusual, with no recognized increase in the risk of malignant melanoma.83
The risk of keloid formation may be more apparent than real because the neck and upper chest, which are the typical areas for operative procedures in these patients, are more likely to have such scarring.98 Premature fine wrinkling of facial skin, similar to that seen in smokers, occurs commonly in women with Turner's syndrome in their late 30s and early 40s. It is not associated with smoking or excessive sun exposure.
Neoplasia
A review of 597 women with Turner's syndrome in the Danish Cytogenetic Register found no increase in the relative risk of cancer, although there were more cases of colon cancer than expected.83 In another review of 400 women, neither colon cancer nor nervous system cancer was increased.99 No history of gonadoblastoma or dysgerminoma was reported in 29 patients with Turner's syndrome and a Y chromosome, but it was not known whether they had undergone prophylactic gonadectomy. One of these patients with a 45,X/46,XY karyotype had adenocarcinoma of a gonadal streak. Two of our 37 patients with Y-chromosome material have had gonadoblastoma. Until better data regarding risk are available, prophylactic gonadectomy is indicated if a Y chromosome is present. Endometrial carcinoma has occurred exclusively in patients who received unopposed estrogen treatment or prolonged treatment with diethylstilbestrol.4
Life Expectancy
Patients with Turner's syndrome appear to have a decreased life expectancy, primarily as a result of complications of heart disease and diabetes.100 In our series of 532 live-born patients, 30 have died, 13 from heart disease (mean age at death, 27.9±25.5 years; range, birth to 80.2 years).4
Summary
Most children with Turner's syndrome are under the care of specialists. It has been proposed that adults should also be followed in multidisciplinary specialty clinics.3 We believe, on the basis of our own experience, that most affected women can best be served by their primary care practitioners, with the use of informed judgment about the need for referral to specialists. Although these women have substantial health concerns, their care for the most part falls under the standard repertoire of primary care, and continued follow-up in specialty care centers may inhibit their integration into society and foster a sense of ill-being. Support groups for patients with Turner's syndrome and their families (listed in the Appendix) can be a source of valuable information.
References
1.-Turner HH. A syndrome of infantilism, congenital webbed neck, and cubitus valgus. Endocrinology 1938;23:566-574. [ISI]
2.-Ford CE, Jones KW, Polani PE, de Almeida JC, Briggs JH. A sex-chromosome anomaly in a case of gonadal dysgenesis (Turner's syndrome). Lancet 1959;1:711-713. [CrossRef][ISI][Medline]
3.-Elsheikh M, Dunger DB, Conway GS, Wass JA. Turner's syndrome in adulthood. Endocr Rev 2002;23:120-140. [Free Full Text]
4.-Sybert VP. Turner syndrome. In: Cassidy SB, Allanson JE, eds. Management of genetic syndromes. New York: Wiley-Liss, 2001:459-84.
5.-Koeberl DD, McGillivray B, Sybert VP. Prenatal diagnosis of 45,X/46,XX mosaicism and 45,X: implications for postnatal outcome. Am J Hum Genet 1995;57:661-666. [ISI][Medline]
6.-Chang HJ, Clark RD, Bachman H. The phenotype of 45,X/46,XY mosaicism: an analysis of 92 prenatally diagnosed cases. Am J Hum Genet 1990;46:156-167. [ISI][Medline]
7.-Gravholt CH, Fedder J, Naeraa RW, Müller J, Fisker S, Christiansen JS. 7.-Occurrence of gonadoblastoma in females with Turner syndrome and Y chromosome material: a population study. J Clin Endocrinol Metab 2000;85:3199-3202. [Free Full Text]

TESTICULO FEMINIZANTE

LA JORNADA,19 FEBRERO 2008
Javier Flores
Testículo feminizante
En los humanos existe una línea básica de desarrollo “femenino”. En años recientes se ha asignado al material genético la mayor importancia en la determinación del sexo. De acuerdo con esto, en términos generales una mujer sería resultado de la presencia de cromosomas sexuales XX, y un hombre de la combinación XY. Pero no es así. Para la formación de los caracteres propiamente masculinos es indispensable la presencia de testículos. Cuando existen fallas en el desarrollo del testículo o en los efectos de las hormonas que este produce, independientemente de las instrucciones genéticas para la determinación del sexo, siempre nacerá una mujer.
Testículo feminizante es el nombre que originalmente se dio a una condición en la que a nivel embrionario, en la combinación genética XY, si bien se desarrollan las gónadas masculinas o testículos, las hormonas que éstos producen –los andrógenos, como la testosterona– carecen de efectos, lo que trae en consecuencia el nacimiento de niñas. En la actualidad se le conoce como síndrome de insensibilidad a los andrógenos (SIA) y se explica por la cancelación parcial o total de los efectos de la testosterona a nivel celular.
De este modo, en la variedad completa del SIA los individuos genéticamente masculinos (XY), con formación de testículos, tienen al nacimiento caracteres sexuales femeninos. En la etapa adulta presentan una morfología externa completamente femenina –con desarrollo mamario y vagina rudimentaria–, a pesar de que carecen de útero y ovarios. En ocasiones esta condición no se descubre hasta la pubertad debido a la ausencia de la menstruación o a causa de su esterilidad, por lo que, si no se detecta a tiempo (y en muchos casos aunque se descubra después del nacimiento), son educadas como mujeres desde la infancia y consideradas como tales hasta la etapa adulta.
Para que una hormona influya sobre las funciones celulares es necesaria la presencia de sitios en la cubierta externa de las células (receptores) que reconozcan la sustancia –en este caso la hormona sexual masculina–, se unan a ella y así se produzcan los cambios en las células, los tejidos, los órganos y en todo el cuerpo que darían como resultado una fisonomía masculina. En el SIA ocurre una mutación genética que altera la estructura de los receptores, lo que los inhabilita para esta función. El resultado es una fisonomía completamente femenina.
Se trata de un síndrome raro, que se presenta con una frecuencia que va de uno en 20 mil nacimientos a uno en 60 mil, de acuerdo con diferentes autores. Pero independientemente de su incidencia, es revelador de la naturaleza humana en el plano sexual. Muestra, al igual que otros síndromes, como el de Swayer, al que me referí aquí la semana pasada, que en los humanos existe una línea básica de desarrollo femenino, independientemente de las instrucciones genéticas para la determinación del sexo.
Si bien las características del síndrome se conocen desde hace muchos años (fue descrito inicialmente por Morris en 1953 como testículo feminizante), hasta hace muy poco se emprendieron estudios serios acerca del papel de género y la conducta sexual de las mujeres con SIA. En otras palabras, si bien existe una fisonomía femenina, la pregunta sería qué pasa a nivel sicológico. En 2003, Hines y sus colaboradores del departamento de sicología de la Universidad de Londres evaluaron en un grupo de 22 mujeres con SIA completo parámetros como la calidad de vida (que incluye la autoestima y el bienestar sicológico general), y las características sicológicas relativas al género (identidad de género, orientación sexual, y el papel y conducta de género en la niñez y la etapa adulta), entre otras pruebas. Los resultados mostraron que no existe diferencia significativa en ninguno de los instrumentos de evaluación aplicados con respecto de un grupo de mujeres consideradas como control (Arch. Sex. Behav. 32(2): 93-101).
De acuerdo con los autores, estos resultados muestran que no se requiere la presencia de dos cromosomas X ni de ovarios para un desarrollo sicológico típicamente femenino, y refuerza la idea de la influencia decisiva del receptor de los andrógenos en el desenvolvimiento de una sicología típicamente masculina.

ANDROGEN INSENSITIVITY SYNDROME; AIS
OMIM# 300068
Alternative titles; symbols TESTICULAR FEMINIZATION SYNDROME; TFMANDRO, GEN RECEPTOR DEFICIENCYAR DEFICIENCY DIHYDROTESTOSTERONE RECEPTOR DEFICIENCY,DHTR DEFICIENCY

Gene map locus Xq11-q12
TEXT
A number sign (#) is used with this entry because the disorder is caused by mutations in the gene for the androgen receptor (AR; 313700).
Adachi et al. (2000) suggested that another form of androgen insensitivity syndrome may be caused by lack of a physiologically indispensable AF-1-specific coactivator crucial to the function of the androgen receptor; see 300274.
DESCRIPTION
The androgen insensitivity syndrome is an X-linked recessive disorder in which affected males have female external genitalia, female breast development, blind vagina, absent uterus and female adnexa, and abdominal or inguinal testes, despite a normal male (2A + XY) karyotype. Partial androgen insensitivity results in hypospadias and micropenis with gynecomastia (Reifenstein syndrome; 312300).
NOMENCLATURE
The androgen insensitivity syndrome was referred to earlier, in both the human and the mouse, as testicular feminization (TFM). The designation androgen insensitivity more accurately reflects the basic pathophysiology and is clearly more satisfactory to patients and their families.
CLINICAL FEATURES
Patients with androgen insensitivity syndrome often come to medical attention because of a presumed inguinal hernia. Many have absent pubic and axillary hair ('hairless pseudofemale'). The hair of the head is luxuriant, without temporal balding. The phenotype is often voluptuously feminine: Netter et al. (1958) reported this disorder in a famous photographic model, Marshall and Harder (1958) reported affected monozygotic twins who worked as airline stewardesses, and Polaillon (1891) described prostitution in an affected person.
In a patient studied by Wilkins (1957), the hair follicles of the axillary and pubic areas, although anatomically normal, were unresponsive to local or parenteral administration of androgens and the beard, voice, and clitoris were similarly unresponsive. This was the first demonstration that the basic defect in cases of the hairless pseudofemale type is end-organ unresponsiveness to androgen, a situation comparable to nephrogenic diabetes insipidus and pseudohypoparathyroidism. (These conditions are analogous to the situation in the Sebright Bantam cock which has a female comb structure despite obvious demonstrations of virility. Albright et al. (1942) misspelled 'Sebright' in their classic article.) It is likely that more than one distinct entity is included in the testicular feminization syndrome. Wilkins stated: 'in about one-third of the cases of male pseudohermaphroditism 'of feminine type' sexual hair has been entirely lacking.'
Morris (1962) called attention to the following case of Gayral et al. (1960): a woman, who was sister, mother, and grandmother of affected males, showed asymmetry in the development of the breasts, body hair, and vulva. The right breast was smaller than the left and there was no pubic hair to the right of the mid-line. She had always had menstrual irregularity but had 3 children, an affected male, a carrier daughter, and a daughter who was the mother of 3 unaffected sons. The findings may be best explained by an X-linked recessive (or incompletely recessive) gene whose effects are to render tissues resistant to male hormone, the patchy changes in the heterozygous female representing the Lyon phenomenon.
According to Wilson (1976), Morris (1953) first described incomplete testicular feminization and concluded that the complete and incomplete forms never occur in the same family. The incomplete syndrome resembles the complete form in respect to female phenotype, bilateral testes and 46,XY karyotype, but differs by clitoral enlargement from birth and virilization at puberty. The abnormality of the external genitalia is characteristic; fusion of the labioscrotal folds occurs for about half of the dorsal portion. Although the degree of masculinization of the external genitalia is variable, most patients are raised as females. In the family described by Lubs et al. (1959), some spermatogenesis was found. There is partial responsiveness to androgen (Winterborn et al., 1970) in this form of the disorder.
It can be difficult to distinguish clinically the incomplete testicular feminization syndrome from pseudovaginal perineoscrotal hypospadias (264600), which is autosomal recessive. Opitz et al. (1972) concluded that the consanguineous family reported by Philip and Trolle (1965) had pseudovaginal perineoscrotal hypoplasia. Boczkowski and Teter (1965) described 3 cases of incomplete testicular feminization among the children of 2 sisters. Wilson (1981) studied 35 families with 1 of the 4 forms of androgen insensitivity: testicular feminization, incomplete testicular feminization, Reifenstein syndrome, or infertile male syndrome. In 31 of the families, he found an abnormality of the androgen receptor: abnormal binding, qualitatively abnormal receptor or decreased amount of receptor. In the other 4, no abnormality of receptor could be demonstrated. Bals-Pratsch et al. (1990) found qualitative and quantitative abnormalities of the androgen receptor in 3 brothers with prepenile scrotum (congenital transposition of the penis), bifid scrotum, scrotal hypospadias, and bilateral undescended testes. In 2 brothers with perineal hypospadias, Batch et al. (1993) found a qualitative androgen binding defect and a point mutation in the AR gene (313700.0020); they suggested that familial hypospadias is part of the phenotypic spectrum of partial androgen sensitivity.
Kaufman et al. (1984) studied an XY patient, with ambiguous genitalia at birth and breast development at puberty, whose cultured fibroblasts showed normal initial formation of low-affinity androgen-receptor complexes but defective transformation of these complexes to a higher affinity state. They presumed that the defect was in the X-linked structural gene for androgen receptor. A qualitative defect of the androgen receptor was demonstrated (Kovacs et al., 1984); although its binding properties were normal, it was unstable on sucrose density gradient centrifugation.
Hughes and Evans (1986) described 2 sibs with classic complete androgen insensitivity syndrome (CAIS) but increased androgen receptor concentrations in genital skin fibroblasts. The steroid-receptor complex appeared to be translocated normally into the nucleus. They concluded that 'the gene coding for the androgen receptor is intact and does not account for the androgen insensitivity.' But is it not possible that the mutation is in the part of the receptor that is concerned with its effects on DNA? Pinsky et al. (1987) described a family in which the proposita and her aunt had partial androgen resistance of a type different from those previously described. Although there was normal maximum binding capacity, there was an increased apparent equilibrium dissociation constant with dihydrotestosterone and 2 synthetic androgens.
Grino et al. (1988) described a family in which gynecomastia and undervirilization occurred in 5 men, 4 of whom had fathered children, in a pedigree pattern consistent with X-linked recessive inheritance. In fibroblasts cultured from genital skin from 2 of the men, the levels of androgen receptor and the affinity of receptor for dihydrotestosterone were normal. However, androgen binding in fibroblast monolayers was thermolabile, upregulation of receptor levels did not occur after prolonged incubation with dihydrotestosterone or methyltrienolone, and dissociation rates at 37 degrees centigrade were increased with the synthetic androgen mibolerone. In addition, in cytosol preparations the androgen receptor protein was unstable. Grino et al. (1988) suggested that this disorder represents the most subtle functional abnormality of androgen receptor characterized to date, since it was compatible with normal male phenotypic development and in some affected men with fertility.
Davies et al. (1997) described 2 patients with complete androgen insensitivity syndrome and mental retardation associated with submicroscopic deletion of the AR gene. They pointed to the report of another patient with associated CAIS and MR. They postulated that the deletion involves, in addition to the AR gene, 1 or more neighboring genes that are implicated in nonspecific MR.
Holterhus et al. (2000) reported a family with 4 affected individuals, 3 brothers (B1-3) and their uncle, displaying strikingly different external genitalia: B1, ambiguous; B2, severe micropenis; B3, slight micropenis; and uncle, micropenis and penoscrotal hypospadias. All had been assigned a male gender. They detected the same mutation in the AR gene (313700.0050) in each subject. Holterhus et al. (2000) demonstrated that the mutant AR could switch its function from subnormal to normal within the physiologic concentration range of testosterone. This was reflected by an excellent response to testosterone therapy in B1, B2, and the uncle. The authors concluded that, taking into account the well documented individual and time-dependent variation in testosterone concentration in early fetal development, their observations illustrated the potential impact of varying ligand concentrations for distinct cases of phenotypic variability in AIS.
Additional abridged information regarding clinical features is available in the clinical synopsis.
BIOCHEMICAL FEATURES
Amrhein et al. (1976) presented evidence for 2 types of testicular feminization: in one, the receptor for dihydrotestosterone (DHT) was deficient; in the other, the receptor (androgen receptor, AR; 313700) was apparently present but the receptor-DHT complex was for some reason ineffective. The second type, 'receptor-positive' cases, included the 3 sibs pictured by McKusick (1964). They displayed some pubic hair. The first type included a patient with the 'hairless female' phenotype, also pictured by McKusick (1964). All were longtime patients of Dr. Lawson Wilkins, and it was in the last patient that he demonstrated unresponsiveness to locally administered androgens.
Griffin (1979) found a qualitative abnormality of androgen receptor, manifested by thermolability, in some cases of testicular feminization. Binding overlapped the normal range at 26 degrees C. It was half-normal at 37 degrees and less than 20% of normal at 42 degrees. Gerli et al. (1979) described a case of complete testicular feminization syndrome in a person with the 47,XXY karyotype. Obviously, nondisjunction occurred in the carrier mother, who was 40 years old. Two sibs and a daughter of each of 2 sisters of the patient also had testicular feminization. Unlike the usual cases, the patient had low plasma testosterone and high gonadotropins. German and Vesell (1966) reported this situation in monozygotic twins. Kaufman et al. (1979) reported 2 'receptor-positive' cases of complete androgen insensitivity. One of these had maternally related affected relatives in 3 successive generations.
Kaufman et al. (1981) suggested that whereas one class of mutation that affects the structural domain of the androgen receptor confers increased dissociability and defective upregulation (a term they coined), a second impairs upregulation only.
AIS results from the incapacity for testosterone and dihydrotestosterone to virilize male embryos and is mainly attributable to molecular defects of the AR gene. In normal males, testosterone and LH rise during the first few months of life, and this physiological surge is commonly used to evaluate the gonadotropic axis at this age. This neonatal surge had not yet been evaluated in detail in newborns with AIS. Bouvattier et al. (2002) sequentially measured plasma testosterone, LH, and FSH during the first 3 months of life in 15 neonates with AIS and AR mutations. A GNRH (152760) and a human CG (see 118860) stimulation test were also performed. Patients were divided into 2 groups with complete or partial AIS (CAIS or PAIS). In patients with PAIS, testosterone levels were in the high-to-normal range at day 30 (18.4 +/- 6.9 nM) and day 60 (12.8 +/- 3.8 nM). In contrast, plasma testosterone values were below the normal range in 9 of 10 patients with CAIS at day 30 (1 +/- 0.3 nM) and day 60 (1.4 +/- 0.7 nM, both P less than 0.004 vs PAIS). Plasma LH values were low in CAIS at day 30 and increased normally in PAIS. Bouvattier et al. (2002) concluded that the postnatal testosterone and LH surge occurs expectedly in neonates with PAIS but is absent in those with CAIS, and that the postnatal testosterone rise requires the receptivity of the hypothalamopituitary axis to testosterone.
To investigate the interaction of androgens with the IGF system, Elmlinger et al. (2001) compared the expression of IGFs and IGFBPs in cultured genital skin fibroblasts from 9 patients with the syndrome of complete androgen insensitivity with that in genital skin fibroblasts from 10 normally virilized males. Complete AIS genital skin fibroblast strains produced significantly lower IGF2 (147470) and IGF2 mRNA than control genital skin fibroblast strains. The production of IGFBP2 (146731) was also decreased in complete AIS genital skin fibroblasts, whereas that of IGFBP3 (146732) did not differ. The authors concluded that in addition to the endocrine actions of IGF1 (147440), IGF2, and IGFBP2, they are also involved as local growth factors in the mediation of androgen action and growth of genital tissues.
Sobel et al. (2006) studied subjects with complete androgen insensitivity and 5-alpha-reductase-2 deficiency (see 264600) to determine the direct effect of androgens on bone mineral density (BMD). In CAI subjects, BMD was significantly decreased in the spine and hip, whereas subjects with 5-alpha-reductase-2 deficiency had normal BMD values. Sobel et al. (2006) concluded that androgens are of direct importance in the development and/or maintenance of BMD and that testosterone and/or low levels of dihydrotestosterone appear to be sufficient for BMD development and/or maintenance.
INHERITANCE
The means for establishing X-linked inheritance include demonstration of linkage with an X chromosome marker, demonstration of lyonization in heterozygous females, and demonstration that the proportion of new mutation cases is one-third rather than one-half (expected of an autosomal dominant). Meyer et al. (1975) found 2 clones of fibroblasts in heterozygous females, one with androgen-binding and one without, thus clinching the X-linkage of this disorder.
CYTOGENETICS
Muller et al. (1990) described an almost 12-year-old black female with testicular feminization and 47,XXY Klinefelter syndrome. Using DNA markers, they demonstrated that the supernumerary X chromosome resulted from maternal nondisjunction during meiosis II. The error at this stage provided the basis for homozygosity of the mutation at the androgen receptor locus.
Xu et al. (2003) described a 3-month-old girl with CAIS in whom the diagnosis was made during elective repair of inguinal hernia, which had been noted shortly after birth. She had a 46,XY karyotype with inversion of the X chromosome with one break disrupting the AR gene. The phenotypically normal 46,XX mother also carried the inversion in one X chromosome; a maternal aunt had CAIS and a 46,inv(X),Y karyotype. At the age of 5 years this aunt had undergone repair of inguinal hernias, at which time testes were identified. She underwent gonadectomy 1 year later because of concerns of potential malignancy. At age 16 years she had primary amenorrhea and a height of 180 cm.
MAPPING
In the most extensively affected kindred known with complete androgen insensitivity, one living in the Dominican Republic, Imperato-McGinley et al. (1990) found linkage to DXS1 and PGK1, localizing the AR gene to an area between Xq11 and Xq13. Linkage between DXS1 and AR showed a peak lod score of 3.2 at theta = 0.06. No recombination was found between PGK1 and AR; peak lod score was 2.9 at theta = 0.0. Although both AR and PGK1 are distal to DXS1, it was not possible to determine the sequence of the 2. Using 3 cDNA probes spanning various parts of the AR gene, they could demonstrate no abnormality in restriction fragment patterns, suggesting that the gene defect is not a deletion but rather a point mutation or a small insertion/deletion.
Also see the mapping section under androgen receptor (AR; 313700).
MOLECULAR GENETICS
See androgen receptor (AR; 313700).
GENOTYPE/PHENOTYPE CORRELATIONS
Boehmer et al. (2001) analyzed the genotype-phenotype relationship in AIS and the occurrence of possible causes of phenotypic variation in families with multiple affected cases. Of 49 index cases with possible AIS identified, 59% had affected relatives. A total of 17 families were studied, 7 families with CAIS (18 patients), 9 families with PAIS (24 patients), and 1 family with female prepubertal phenotypes (2 patients). No phenotypic variation was observed in families with CAIS. However, phenotypic variation was observed in 1 of 3 families with PAIS resulting in different sex of rearing and differences in requirement of reconstructive surgery. Intrafamilial phenotypic variation was observed for mutations R846H (313700.0040) and M771I (313700.0039). Patients with a functional complete defective AR had some pubic hair, Tanner stage P2, and vestigial wolffian duct derivatives despite absence of AR expression. Vaginal length was functional in most but not all CAIS patients. Boehmer et al. (2001) concluded that while phenotypic variation was absent in families with CAIS, distinct phenotypic variation was observed relatively frequent in families with partial AIS.
PATHOGENESIS
French et al. (1966) found that testosterone failed to affect the urinary excretion of nitrogen, phosphorus and citric acid when given in a dosage much greater than that which in controls decreased excretion of all three. Plasma estrogen levels were the same as those observed in the normal female. Leydig cell stimulation to estrogen production occurs probably because of failure of the feedback repression of the pituitary which shares the unresponsiveness to testosterone. Southern and Saito (1961) showed normal testosterone levels in this disorder.
CLINICAL MANAGEMENT
Ong et al. (1999) identified a met807-to-thr mutation (313700.0044) in the AR gene in a 46,XY infant with female-appearing genitalia. An AR construct bearing the met807-to-thr mutation, when expressed in COS-7 and HeLa cells, did not bind to its natural ligand testosterone at its upper physiologic range of 3 nM/L but bound dihydrotestosterone to near normal levels at the same concentration. The transactivation function of the receptor was reduced to 15% of normal at physiologic doses of the hormone. However, DHT was able to induce receptor transactivity 10-fold more efficiently than testosterone, in parallel with androgen-binding assays. Intramuscular injections of increasingly higher doses of depo-testosterone, though elevating serum testosterone level, did not result in significant development of the infant's male external genitalia. Subsequently, he was treated with a DHT gel, applied topically to the periscrotal region 3 times a day, for 5 weeks. Serum DHT rose from 1.8 nmol/L to 8.8 nmol/L and resulted in improved male genital development. The authors concluded that in vitro functional assays can help identify the subset of patients with ambiguous genitalia who could respond well to androgen therapy, providing them an option to be reared in accordance with their chromosomal sex.
Wisniewski et al. (2000) assessed by questionnaire and medical examination the physical and psychosexual status of 14 women with documented complete androgen insensitivity syndrome (CAIS). They determined participant knowledge of CAIS as well as opinion of medical and surgical treatment. As a whole, secondary sexual development of these women was satisfactory, as judged by both participants and physicians. In general, most women were satisfied with their psychosexual development and sexual function. All of the women who participated were satisfied with having been raised as females, and none desired a gender reassignment. Although not perfect, the medical, surgical, and psychosexual outcomes for women with CAIS were satisfactory; however, specific ways for improving long-term treatment of this population were identified.
POPULATION GENETICS
Mainly using data on the frequency of inguinal hernia in females, Jagiello and Atwell (1962) estimated the frequency of testicular feminization as being about 1 in 65,000 males.
Edwards et al. (1992) demonstrated that the distribution of the number of CAG repeats in exon 1 of the AR gene was lowest in African Americans, intermediate in non-Hispanic whites, and highest in Asians. The distribution of allele size was bimodal in African Americans, and only in African Americans was there a deviation from Hardy-Weinberg equilibrium. Irvine et al. (1995) studied the distribution of the CAG and GC repeats (microsatellites) in exon 1 of the AR gene in African Americans, non-Hispanic whites, and Asians (Japanese and Chinese) and confirmed the findings of Edwards et al. (1992). The frequency of prostate cancer (176807) in the 3 racial groups is inversely proportional to the length of the repeats. One of the critical functions of the product of the AR gene is to activate the expression of target genes. This transactivation activity resides in the N-terminal domain of the protein which is encoded in exon 1 which contains the polymorphic repeats. The smaller size of the CAG repeat is associated with a higher level of receptor transactivation function, thereby possibly resulting in a higher risk of prostate cancer. Irvine et al. (1995) noted that Schoenberg et al. (1994) had observed a somatic mutation resulting in a contraction of the CAG repeat from 24 to 18 in an adenocarcinoma prostate and the effects of the shorter allele were implicated in the development of the tumor.
Based on patients with molecular proof of the diagnosis in a nationwide study in the Netherlands and previous estimates from the Danish patient registry, Boehmer et al. (2001) estimated that the minimal incidence of AIS is 1:99,000.
ANIMAL MODEL
Lyon and Hawkes (1970) described a homologous phenotype in the mouse and showed that it is genetic, the Tfm locus being situated in the middle of the X chromosome. Ohno and Lyon (1970) showed that in these mice certain enzymes of the mouse kidney, e.g., alcohol dehydrogenase, are not inducible by testosterone as is usually possible. They postulated that the Tfm locus is a repressive regulatory locus controlling many testosterone inducible enzymes. In affected hemizygotes all these enzymes become noninducible. According to their suggestion, this is a regulator mutation like the noninducible mutation in the lac-repressor locus of E. coli as elucidated by Jacob and Monod (1963). Bardin et al. (1970) described studies of the pseudohermaphroditic rat which seems to have a disorder analogous to testicular feminization. Androgen-dependent differentiation is absent. Defective formation of dihydrotestosterone was apparently not the explanation. Goldstein and Wilson (1972) studied the Tfm mouse and showed, by giving dihydrotestosterone to pregnant mothers, that there is resistance to androgen-mediated sexual differentiation in embryos. Low serum testosterone and low production of testosterone in adult Tfm testis of the mouse were features different from those in man, but were considered by them as secondary to the defect in differentiation. They showed deficient binding of testosterone in the nuclei of the submaxillary gland of these adult Tfm animals, but again this may be the result of incomplete differentiation of an androgen-sensitive cell line.
Bullock and Bardin (1972) concluded that androgen-binding proteins are absent from the cytosol of preputial gland of Tfm rats and from the kidney of Tfm mice. Testicular feminization rats, despite female external sexual development, show masculine sexual behavior and little feminine sexual behavior. In the Tfm mouse, Charest et al. (1991) demonstrated a single base deletion in the N-terminal domain of the androgen receptor, resulting in a frameshift mutation. Gaspar et al. (1991) independently demonstrated the same abnormality. They found no structural aberration in the coding region of the messenger by a series of RNase-protection assays. However, cell-free translation of RNAs transcribed in vitro from enzymatically amplified overlapping segments of exon 1 demonstrated a truncated receptor protein. Sequence analysis showed deletion of a single nucleotide in the hexacytidine stretch at position 1107-1112 altering the reading frame of the messenger and introducing 41 missense amino acids before the premature termination codon at position 1235-1237.
In female mice heterozygous for the Tfm gene, Takeda et al. (1987) demonstrated mosaicism in 2 androgen target tissues by steroid autoradiographic techniques, thus documenting X-linked inheritance. See also Takeda et al. (1987).
HISTORY
The variety of sex anomaly described in this entry has been of relatively long interest to geneticists, largely through the publication of Pettersson and Bonnier (1937), who concluded that the affected persons are genetic males. Dieffenbach (1912), an American geneticist, had pointed out the hereditary pattern. Morris (1953), in a classic paper, first used the term testicular feminization.
Miller (1961) considered 'feminizing labial testes' of the type described by Lubs et al. (1959) to be a separate form of male pseudohermaphroditism. However, Wilson et al. (1984) described well-studied cases that indicated that the Lubs syndrome (Lubs et al., 1959), like classic testicular feminization, is due to mutation in the androgen receptor. The patients were first cousins; their mothers were sisters.
REFERENCES
1. Adachi, K.; Kano, M. :
Adenyl cyclase in human hair follicles: its inhibition by dihydrotestosterone. Biochem. Biophys. Res. Commun. 41: 884-890, 1970. PubMed ID : 4320069
2. Adachi, M.; Takayanagi, R.; Tomura, A.; Imasaki, K.; Kato, S.; Goto, K.; Yanase, T.; Ikuyama, S.; Nawata, H. :
Androgen-insensitivity syndrome as a possible coactivator disease. New Eng. J. Med. 343: 856-862, 2000. PubMed ID : 10995865
3. Albright, F.; Burnett, C. H.; Smith, P. H.; Parson, W. :
Pseudo-hypoparathyroidism, an example of 'Seabright-Bantam syndrome': report of 3 cases. Endocrinology 30: 922-932, 1942.
4. Amrhein, J. A.; Meyer, W. J., III; Jones, H. W., Jr.; Migeon, C. J. :
Androgen insensitivity in man: evidence for genetic heterogeneity. Proc. Nat. Acad. Sci. 73: 891-894, 1976. PubMed ID : 176660
5. Bals-Pratsch, M.; Schweikert, H.-U.; Nieschlag, E. :
Androgen receptor disorder in three brothers with bifid prepenile scrotum and hypospadias. Acta Endocr. 123: 271-276, 1990. PubMed ID : 2146851

DETECCION TEMPRANA DE GENES DEL CANCER

Systematic Review: Gene Expression Profiling Assays in Early-Stage Breast Cancer

Luigi Marchionni, MD, PhD; Renee F. Wilson, MSc; Antonio C. Wolff, MD; Spyridon Marinopoulos, MD, MBA; Giovanni Parmigiani, PhD; Eric B. Bass, MD, MPH; and Steven N. Goodman, MD, MHS, PhD
4 March 2008 Volume 148 Issue 5 ANNALS INTERNAL MED.
Background: Three gene expression–based prognostic breast cancer tests have been licensed for use.
Purpose: To summarize evidence on the validity and utility of 3 gene expression–based prognostic breast cancer tests: Oncotype DX (Genomic Health, Redwood City, California), MammaPrint (Agendia BV, Amsterdam, the Netherlands), and H/I (AvariaDX, Carlsbad, California).
Data Sources: MEDLINE, EMBASE, and Cochrane databases (from 1990 through January 2007), Web sites of the test manufacturer, and discussion with the manufacturers.
Study Selection: Original data studies published in English that addressed prognostic accuracy and discrimination or treatment benefit prediction of any of the 3 tests in women with breast cancer.
Data Extraction: Information was extracted about the clinical characteristics of the study population (particularly clinical and therapeutic homogeneity), tumor characteristics, and whether the marketed test or underlying signature was evaluated.
Data Synthesis: The tests are based on distinct gene lists, using 2 different technologies. Overall, the body of evidence showed that this new generation of tests may improve prognostic and therapeutic prediction, but the tests are at different stages of development. Evidence shows that the tests offer clinically relevant, improved risk stratification over standard predictors. Oncotype DX has the strongest evidence, closely followed by MammaPrint and the H/I test (which is still maturing).
Limitations: For all tests, the relationship of predicted–observed risk in different populations needs further study, in addition to their incremental contribution over conventional predictors, optimal implementation, and relevance to patients receiving current therapies.
Conclusion: Gene expression technologies show great promise to improve predictions of prognosis and treatment benefit for women with early-stage breast cancer. More information is needed on the extent of improvement in prediction, in whom the tests should be used, and how test results are best incorporated into decision making about breast cancer treatment.
Currently, 3 commercially available prognostic breast cancer tests based on gene expression (see Glossary) technology are available: Oncotype DX (Genomic Health, Redwood City, California), MammaPrint (Agendia BV, Amsterdam, the Netherlands), and H/I (AvariaDX, Carlsbad, California). Although measurement of gene expression is now a core research method, these commercial assays represent the first introduction of these technologies into clinical application.
Gene expression is the technical term to describe how active a particular gene is—that is, how many times it is expressed, or transcribed, to produce the protein it encodes (Figure 1). The transcription (see Glossary) of the gene's DNA into messenger RNA (mRNA) is the first step in this process; modern molecular biological tools allow this activity to be measured by counting the number of mRNA molecules within a given cell type or tissue. Because the mRNA molecule is translated within the ribosome to produce a complete protein, counting mRNA transcripts provides an estimate of the number of corresponding proteins. High-throughput technologies, such as DNA microarray (see Glossary) and real-time reverse transcriptase polymerase chain reaction (RT-PCR) (see Glossary), allow simultaneous counting of several gene transcriptions (up to tens of thousands). This creates a snapshot of a tissue's global gene activity, called the transcriptome.
Figure 1. (A) Technologies enabling high throughput gene expression analysis. Breast cancer tumors are sampled at the treatment location and shipped to the central laboratory doing the assay, where pathologic review is done to assess cancer cell contents, followed by RNA preparation and integrity evaluation. Suitable samples are used to quantify RNA levels, thus assessing gene expression. When a gene is expressed, the transcription complex copies its DNA sequence into complementary RNA transcripts that are translated into proteins. High-throughput gene expression analysis aims at the quantification of messenger RNA (mRNA) populations in a given tissue. (B) Deoxyribonucleic acid microarray is the molecular biology technique enabling gene expression analysis in the MammaPrint assay. Ribonucleic acid is labeled with fluorescent dye and hybridized against thousands of different nucleotide sequences corresponding to different genes and arrayed on a solid surface (that is, a modified microscope glass slide). On hybridization, fluorescence emitted by single locations on the microarray is used to estimate gene expression levels. In MammaPrint a 2-color design is used, and RNA expression is estimated as a relative ratio between the sample and a reference RNA. For each patient, triplicate measurements are obtained from 2 microarray inverting the labeling scheme. (C) Real-time reverse transcriptase polymerase chain reaction is the enabling technology to assess gene expression in Oncotype DX and H/I tests. This technique is based on reverse transcription (see Glossary) of a specific mRNA into the complementary DNA molecule, which is used as a template in a polymerase chain reaction. The production of double-stranded DNA is accompanied by emission of light, which is recorded throughout the process and correlates to the amount of DNA that is produced. The input RNA is more concentrated in the initial reverse transcription reaction, and the earlier light is emitted during polymerase chain reaction, thus different levels of gene expression correspond to delayed light emission measurement (earlier vs. later) in the polymerase chain reaction step. (D) Gene-expression levels are translated by different mathematical transformation into indexes predicting disease recurrence (D).
Gene expression measurements have been used to develop new biological concepts, refine disease classification, improve diagnostic and prognostic accuracy, and identify new molecular targets for drugs, especially in cancer research (1–9). Results are commonly reported in the form of a list of genes that are differentially expressed between normal and diseased patients or that correlate with different prognoses or phenotypes. These lists are called gene expression profiles or signatures(see Glossary).
"Breast cancer" is increasingly understood as an umbrella designation for various tumor subtypes that differ in their prognoses and responses to therapy. An important decision for many patients with early-stage breast cancer, especially patients who have tumors that express hormone receptors and will be treated with antiestrogen therapy, is whether they should also be treated with systemic chemotherapy. Although adjuvant chemotherapy is frequently recommended in this setting, many women would have remained recurrence-free at 10 years without it, especially those with small, estrogen receptor–positive tumors, without axillary nodal involvement. Patients and their physicians must weigh the possible benefit of chemotherapy in reducing this risk against its toxicity and other attendant costs.
Practicing oncologists frequently base their decisions about therapy on prognostic clinical algorithms that include demographic data; tumor stage; and other tumor characteristics, such as grade and estrogen receptor expression. These conventional combination predictors include the National Institutes of Health (NIH) Consensus Development criteria (10, 11); the St. Gallen expert opinion criteria (12, 13); the National Comprehensive Cancer Network guideline (14–16); and a Web-based algorithm, Adjuvant! Online (17, 18). Gene expression profiling has been proposed to potentially augment or replace these prognostic tools.
Oncotype DX is based on a 21-gene profile developed by Paik and colleagues (19), MammaPrint is based on a 70-gene prognostic signature developed by van't Veer and colleagues (8), and the H/I assay is based on a 2-gene ratio signature (HOXB13–IL17BR developed by Ma and colleagues (20). The gene sets on which these tests are based almost do not overlap: The 21-gene and the 70-gene expression signatures that form the basis of the Oncotype DX and MammaPrint assays, respectively, share only 1 gene in common. Two technologies are used to determine gene expression: real-time RT-PCR (Oncotype DX and H/I) and DNA microarray (MammaPrint). All 3 tests use pathologic review of specimens to check tumor content and evaluate RNA preparation and quality. The 2 RT-PCR–based assays (Oncotype DX and H/I) are done in formalin-fixed, paraffin-embedded tumor tissues, whereas fresh unfixed tumor tissue is required for MammaPrint. We review evidence on the prognostic accuracy of these 3 tests and their ability to predict treatment benefit.
Methods
Data Sources
On 9 January 2007, we searched the MEDLINE and EMBASE databases by using Medical Subject Headings and other terms relevant to breast cancer, gene expression profiling, and Oncotype DX or MammaPrint. On 7 February 2007, we searched the Cochrane database, including Cochrane Reviews, CENTRAL, and CINAHL. We supplemented this search by updating searches in MEDLINE as well as by hand-searching added publications that appeared after the initial search (January 2007 to July 2007) and studies related to the H/I test. The test manufacturers were also asked to provide any published or unpublished data relating to our study questions. Searches were limited to publications in English.
Study Selection
Two investigators independently reviewed titles and abstracts to identify original data studies that involved the use of any of the 3 assays in women with breast cancer.
Data Extraction
We extracted and double-checked information on the clinical characteristics of the study population, tumor characteristics, and whether the marketed test or underlying signature was evaluated. To assess the quality of studies, we applied (where appropriate) the general principles of the REMARK (REporting recommendations for tumor MARKer prognostic studies) (21, 22) and Standards for Reporting of Diagnostic Accuracy (23, 24) guidelines.
We synthesized data on the ability of a test to accurately predict recurrence risk (clinical validity) and treatment benefit (clinical utility). We distinguished between gene expression signatures and the gene expression–based marketed tests. The gene signature is the collection of genes whose expression levels are measured in a given test, together with the algorithm that combines those levels into a prognostic index. A gene signature can be measured by using various technologies (RT-PCR or complementary DNA [cDNA] array) and procedures (for example, different reagents, controls, sample acquisition, preparation, and transport procedures), which may not be identical to those used in the marketed test.
We report limited information on the technical performance characteristics of the tests, sometimes called analytic validity. The analytic validity of a test usually is assessed by determining how observed measurements differ from standard reference values. However, no reference standard exists for gene expression measurements outside of the technologies used for these tests. However, because analytic validity affects predictive ability, our assessment of predictive ability successfully incorporates the effect of less-than-perfect analytic validity. In the full evidence report (available at www.ahrq.gov/clinic/epcindex.htm), we summarize data on the reproducibility of a test when applied repeatedly to the same patient or when repeated over time, as well as variability as a function of tumor sampling and handling, specimen preparation, and biological variation within tumor samples. Appendix Tables 1, 2, and 3 show the evidence summary
Role of the Funding Source
The Agency for Healthcare Research and Quality and the Centers of Disease Control and Prevention's Evaluation of Genomic Applications in Practice and Prevention program helped formulate the initial study questions but did not participate in the literature search, determination of study eligibility criteria, data analysis, or interpretation.
Results
shows the number of studies considered at each phase of title, abstract, and article review. The final set of 26 studies was heterogeneous in focus and quality. Few reports addressed technical aspects of the tests. Ten reports focused on prognostic prediction. Only 1 study, involving Oncotype DX, examined the prediction of treatment benefit. Most of the published evidence available for Oncotype DX was conducted with the marketed assay. The evidence relevant to MammaPrint was a mix of studies of the underlying signature and of the marketed test. Only 1 study used the marketed version of H/I (41), and it was not clear whether the laboratory doing the assay in that report was the same as the one with current rights to do the test. All other studies relevant to H/I examined the underlying 2-gene signature, using different measurement techniques and algorithms than the ones implemented in the marketed- test.
Analytic Validity and Variability
We found limited evidence about the laboratory procedures used for Oncotype DX (26, 27) and MammaPrint (34, 35), including information about their reproducibility. Such evidence was reported in methodological studies (26–35) and in clinical studies that focused on predictive validity (19, 30). In the case of MammaPrint, results from different laboratories depended on RNA labeling protocols (34), suggesting that MammaPrint results may not be similar if done in different laboratories (only 1 laboratory currently offers the test). The overall proportion of samples that were successfully tested with the various methods ranged from 67.7% to 98.9% for Oncotype DX and 67.7% to 80.9% for MammaPrint (36) (Appendix Tables 1 and 2). No reports investigated the reproducibility of H/I; Ma and colleagues (41) reported a success rate of 98%.
Predicting Disease Outcomes
Oncotype DX
Oncotype DX was developed on the basis of a prospectively chosen 250-candidate gene set, which was then measured on 447 patients with breast cancer who were treated in 1 of 3 completed randomized trials with long-term follow-up. From these 250 genes, 21 genes (16 cancer-related and 5 references) were chosen to predict 10-year breast cancer recurrence. The expression levels of these genes are measured by using RT-PCR combined with a published quantitative algorithm to produce a number between 0 and 100, which is the recurrence score. In this review, "recurrence score" indicates the numeric value generated from Oncotype DX (19). The recurrence score is categorized into 3 risk strata: low (score <18),>18 but <30),>19, 30, 31, 25). Paik and colleagues (19) studied 668 women in a randomized, controlled trial conducted by the National Surgical Adjuvant Breast and Bowel Project (NSABP) (Appendix Table 1). The parent study (the NSABP B-14 trial), which enrolled patients from 1982 to 1988, examined the effect of tamoxifen therapy versus placebo in women with lymph node–negative, estrogen receptor–positive, early-stage breast cancer (42–45). The recurrence score algorithm and risk categories for the group treated with tamoxifen were prespecified on the basis of the development studies. Stratification into the 3 risk categories yielded univariate actuarial 10-year recurrence risks of 7% (low), 14% (intermediate), and 31% (high) (P < href="http://www.annals.org/cgi/content/full/0000605-200803040-00208v1#TA1">Appendix Table 1). The recurrence score was the strongest predictor among all traditional risk factors, with an adjusted hazard ratio (HR) of 2.8 (CI, 1.7 to 4.6) for a 50-point change in the risk strata.
Glas and colleagues (35) examined the clinical performance of the Oncotype DX assay to predict breast cancer death at 10 years in a community-based population of lymph node–negative, estrogen receptor–positive patients treated with tamoxifen. Two hundred twenty case patients (dead) and 570 matched control patients (alive) were selected, and 165 case patients with an estrogen receptor–positive status and 55 case patients who received tamoxifen treatment (among estrogen receptor–positive patients) formed the final study sample. For patients treated with tamoxifen, results paralleled those of the NSABP B-14 trial (which examined recurrence): The probability of death at 10 years was 2.8%, 11%, and 16% in the low-, medium-, and high-risk groups, respectively. These rates were about 3 percentage points higher among patients not treated with tamoxifen. Prognostic value persisted after stratification by tumor grade and disease stage. The continuous risk strata also showed a relationship with mortality risk in 52 estrogen receptor–negative patients after adjustment for tumor grade and disease stage (relative risk [RR], 1.4 per 10-unit increase in risk [CI, 1.04 to 2.0]). Another study (36) showed no predictive value of the recurrence score in a small population of patients who received neither tamoxifen nor chemotherapy. However, worse tumor grade predicted better prognosis in that study, suggesting that the results were not reliable. A fourth paper on the signature alone, in which the main purpose was to contrast the different tests (33) (described below in Comparison of Signatures).
No published study showed how the recurrence score reclassified patients into different risk strata after initial classification by conventional predictors. However, in 2004, Paik and colleagues (46) presented further information in poster form. They found that the recurrence score had predictive power beyond that of the St. Gallen or National Comprehensive Cancer Network risk stratification guidelines sufficient to change some patient decisions about chemotherapy (The St. Gallen test did not include human epidermal growth factor receptor 2 at that time, which is included in the recurrence score). On the basis of the 2004 National Comprehensive Cancer Network guidelines, the study indicated that about half of the 92% of patients who were in the high-risk National Comprehensive Cancer Network category were reclassified as low-risk by the recurrence score, with a 10-year relapse risk of 7% (CI, 4% to 11%); this is similar to the risk seen in the low-risk recurrence score group without the National Comprehensive Cancer Network information. The same information for Adjuvant! Online was part of an oral presentation in 2005 (Table 1) (47). Compared with the Adjuvant! Online criteria, roughly 40% of women assessed to be at high risk (22% relapse) were reclassified into a group with an observed 8% risk if they had a low recurrence score. With Adjuvant! Online, 39% of women classified as high risk (31% recurrence) had a 9% recurrence risk after a low recurrence score. These findings showed that the greatest contribution of the test is probably the reclassification of patients from high to low risk (that is, reducing the number of patients who might unnecessarily undergo chemotherapy) and combining this test with conventional predictors yields the most information.
MammaPrint
MammaPrint is based on the 70-gene signature derived from an initially unselected set of more than 25 000 candidate genes on a cDNA array. The test was developed in 2002 at the Netherlands Cancer Institute using 78 lymph node–negative patients younger than age 55 years who did not carry a breast cancer gene mutation and who had tumors that were less than 5 cm in diameter (8). The end point for this training set was 5-year distant recurrence. Patients are classified by calculating the correlation coefficient between a patient's expression levels of the 70 genes and an average good-prognosis expression profile. If the correlation coefficient exceeds 0.4, the patient is classified as having a good prognosis; if less, they are classified as having a poor prognosis.
van de Vijver and coworkers (9) validated this signature in a series of 295 consecutive patients with stage I or II breast cancer and small tumors (<5>35) reanalyzed 145 patients from van de Vijver and coworkers' (9) cohort and all 78 patients from the training set by using the marketed MammaPrint assay instead of the signature (35). A different reference RNA and different quantification method were used. Although odds ratios (ORs) and HRs were similar to those found in the earlier studies, approximately 10% (7 of 78) of patients were placed into different risk categories, most of which had borderline correlations.
The MammaPrint test was validated in a multicenter European study of 302 patients not treated with chemotherapy or tamoxifen and provided prognostic information beyond that of Adjuvant! Online (36). Frozen tumor specimens from node–negative patients younger than age 60 years who did not receive systemic adjuvant chemotherapy were tested. Ninety patients were estrogen receptor–negative, and none of the estrogen receptor–positive patients received tamoxifen. The median follow-up was 13.6 years, and the overall rate of distant metastasis was 25%. The area under the receiver-operating characteristic curves indicated that both methods had similarly modest discriminatory power in absolute terms (0.68 for MammaPrint and 0.66 for Adjuvant! Online), but MammaPrint provided better reclassification of patients in risk groups (Table 2). Hazard ratio estimates between high- and low-risk categories for distant recurrence in van de Vijver and colleagues' study were substantially higher than those in the validation study (unadjusted HR, > 15 vs. 2.3, respectively; adjusted HR, 4.6 vs. 2.1). Compared with van de Vijver and colleagues' (9) study, the validation cohort was observed for a longer period (median, 13.6 vs. 6.7 years), included older women, and excluded patients who received adjuvant therapy. The validation study also found that HRs for all end points decreased steadily with an artificial increase in censoring time from 5 to 10 years.
Ma and colleagues (20) identified the 2 genes that are the basis for the H/I test by screening 22 000 genes in 60 patients with estrogen receptor–positive, lymph node–positive or negative breast cancer treated with tamoxifen. High expression of HOXB13 predicted recurrence, and high expression of IL17BR predicted nonrecurrence; therefore, a higher ratio of the 2 genes strongly predicted recurrence in this training set (interquartile OR, 10.2; adjusted OR, 10.2).
Reid and colleagues (37) examined 58 tamoxifen-treated patients with estrogen receptor–positive breast cancer whose disease was more advanced disease than in Ma colleagues' sample (48). No relationship between the expression of these genes and distant relapse was observed in these patients or in an additional 99 patients derived from a previously studied cohort (5) that the authors investigated after the initial negative result. In 2006, Goetz and colleagues (38) analyzed 206 estrogen receptor–positive patients treated in the tamoxifen-only arm of a phase III randomized trial. Expression values were normalized by using a different approach than that used by Ma and colleagues (20), as well as different cutoff points for the ratio that best predicted relapse-free survival, disease-free survival, and overall survival were calculated. The ratio had modest value for outcome prediction in the entire cohort, with cross-validated HRs near 1.5 and P values around 0.05, but this predictive ability was restricted to the node–negative subset of patients.
In a large validation study, Ma and colleagues (41) examined a consecutive series of 852 patients with stage I or II breast cancer at a median follow-up of 6.8 years. The investigators used a slightly different method from what previously used (20) to combine and normalize the expression of the 2 genes into an index that is now the basis of the H/I assay. In a stratified analysis, the HOXB13–IL17BR ratio was predictive only in patients with node–negative, estrogen receptor–positive disease. The investigators optimized the threshold (maximizing the HR) differently in patients treated with tamoxifen and those who were not. The adjusted HR incorporating other risk factors was 3.9, regardless of tamoxifen treatment. Classification probabilities were not presented, and the incremental value of the HOXB13–IL17BR ratio compared with conventional combined predictors was not reported, although some components of those predictors were included in the multivariable analyses.
Jansen and associates (39) evaluated the ability of the HOXB13–IL17BR ratio to predict disease-free survival in 1252 patients with breast cancer who had undergone various surgical treatments. In this group, 73% of tumors were estrogen receptor–positive; 52% of patients were lymph node–positive, 14% were treated with tamoxifen, 17% received chemotherapy; and 55% of patients received tamoxifen or chemotherapy after relapse. Jansen and associates (39) used different populations, protocols, normalization strategy, and ratio thresholds than Ma and colleagues (41). The overall relapse rate was high at 51% after a median of 6 years follow-up. The HOXB13–IL17BR ratio was examined in 468 patients with lymph node–negative, estrogen receptor–positive disease who did not receive adjuvant systemic chemotherapy. The ratio was associated with poor disease-free survival in a multivariable model (HR, 1.6; P = 0.02) and poor overall survival (HR not reported; P < href="http://www.annals.org/cgi/content/full/0000605-200803040-00208v1#TA3">Appendix Table 3). The ratio was not compared with conventional combination risk indices, and classification probabilities for the models with and without the ratio were not provided.
Jerevall and coworkers (40) investigated whether the HOXB13–IL17BR ratio predicted a differential benefit between 264 patients with postmenopausal breast cancer who received tamoxifen for 2 and 5 years and 93 premenopausal patients who did not receive systemic therapy. Seventy-two percent of patients had lymph node–positive disease and 74% had estrogen receptor–positive disease (Appendix Table 3). The authors dichotomized the HOXB13–IL17BR ratio at the median, which differed from the approach used by Ma and colleagues (41). Jerevall and coworkers (40) concluded that IL17BR might be an independent prognostic factor in breast cancer and suggested that HOXB13 may be correlated with tamoxifen resistance. However, the HOXB13–IL17BR ratio had no prognostic value in postmenopausal patients with estrogen receptor–negative disease. Neither the patient profile nor the methods of calculation of the ratio were identical to those used in previous studies, and the results differed from previous reports because the ratio HOXB13–IL17BR predicted worse outcomes in patients with lymph node–positive disease.
Comparison of Signatures
Fan and colleagues (33) used the same data set evaluate both the agreement between gene expression tests and other predictors and the individual performance of the tests. The Oncotype DX risk strata and the HOXB13–IL17BR ratio were estimated from microarray gene expression data (that is, not RT-PCR) and thus were not obtained according to the protocols and methods used in the marketed assays. These data are therefore described as derived scores. Fan and colleagues (33) used the same 295 samples from patients with stage I or II breast cancer that had been used to develop the 70-gene signature (9). Therefore, Fan and colleagues' (33) comparison would be expected to favor the 70-gene profile over the derived recurrence score and HOXB13–IL17BR ratio.
The 70-gene signature and the derived recurrence score predicted overall survival and disease-free survival, but the derived HOXB13–IL17BR ratio did not predict either (HR, about 1); however, measurement of the derived 2-gene ratio may have been flawed (49). The intermediate- and high-risk groups, as defined by the derived recurrence score, were combined and compared with the poor-prognosis group defined by the 70-gene signature. The agreement between MammaPrint and the derived recurrence score was 81% (239 of 295). All analyses were repeated for the 225 patients in the estrogen receptor–positive subset, with qualitatively similar results. Good correlation between predictions was found, which was of interest because classification was obtained by using different gene sets. The degree of prediction over and above standard combined clinical stratifiers was not clear, and a reclassification analysis of patients was not done.
Predicting Treatment Response
The ability of the Oncotype DX) test to predict chemotherapy benefit was investigated in patients from the NSABP-20 trial (50). In this study, Paik and colleagues (50) examined 10-year, distant recurrence-free survival in 651 patients with estrogen receptor–positive, lymph node–negative disease who were randomly assigned to receive tamoxifen alone or tamoxifen with chemotherapy. An overall benefit was seen for chemotherapy, but when the data were stratified by risk group, the benefit was restricted to patients with a high recurrence score (RR, 0.26 [CI, 0.13 to 0.53]), a finding that persisted in multivariate analyses. However, even though no benefit was seen in the low recurrence-score group, the point estimate had very wide CIs; a clinically relevant benefit could therefore not be excluded.
Two studies examined whether the recurrence score predicted pathologic response in patients receiving preoperative systemic therapy (28, 29). Neither study was done at the laboratory offering the Oncotype DX assay (Appendix Table 1). One study found that the recurrence score predicted complete response (28), whereas the other study (29) found no such relationship Finally, Chang and colleagues (32) assessed chemotherapy response prediction in 12 patients with complete clinical response among 72 women enrolled in phase II studies of docetaxel and found that a high recurrence score was associated with complete response (P = 0.008). When the recurrence score was used as a continuous variable, a 14-unit increase in the score (the difference between the low- and high-risk groups, as defined by the standard thresholds) insubstantially predicted a clinical complete response (OR, 1.7 [CI, 1.15 to 2.60]).
No study investigated the ability of MammaPrint to predict treatment response, but 1 study reported that the HOXB13–IL17BR ratio could predict whether 5 years of tamoxifen therapy would provide survival benefit over 2 years of tamoxifen treatment in estrogen receptor–positive patients (40).
Discussion
This body of evidence on the 3 marketed gene expression tests for breast cancer prognosis shows that these tests have considerable potential for improving prognostic and therapeutic prediction. It also provides valuable lessons about the complexity in evaluating such tests.
Because the role of the genes included in these tests in determining prognosis is not completely understood, it is often unclear which clinical or tumor characteristics are being measured. Intrinsic tumor aggressiveness, ability to metastasize, and responsiveness to treatment (hormonal, radiation, or chemotherapy)—each of which might involve different genes—can be used to determine prognosis. However, the measure being assessed is that a particular study must sometimes be inferred from the treatment, tumor, and clinical characteristics of the study population. Results from populations that are clinically and therapeutically heterogeneous may not be optimal in determining the prognosis or risk for a particular woman. In addition, survival was defined in the studies as disease-free, distant recurrence-free, or overall and over 5 or 10 (or more) years, and prediction strength varied considerably depending on what the test was optimized for. Finally, performance of the underlying gene signature is often close but not identical to the marketed test, because many test procedures, including pretest sample preparation and transport, can differ. It is therefore critical to pay close attention to the test description, population, and end points for each study to understand which studies are mutually supportive, which are adding qualitatively different information, and to whom they apply.
Despite the clinical novelty of these tests, their developmental pathway must continue by following the same principles and procedures as those for any multivariate clinical prediction rule. These are outlined in detail in the clinical literature (51, 52) and have been agreed on in reporting guidelines (22) and articulated with respect to expression-based predictors in various review articles (53–55).
Each of the 3 marketed tests is at a different point in the developmental pathway. Almost all studies of Oncotype DX have used the marketed test as opposed to the signature, whereas the evidence on MammaPrint comes from studies examining the signature and the assay (only the large multicenter validation by Buyse and colleagues (36) used the marketed assay). The study that compared the results of the marketed MammaPrint test versus its signature on the same samples showed that about 10% of the patients were placed into different risk groups when the marketed test was used. Almost all of the studies based on H/I calculated or implemented the test in subtly different ways, and only 1 seemed to use the marketed H/I test (48).
In terms of the clinical and therapeutic homogeneity of the underlying populations, Oncotype DX focused on a narrower, more clinically and therapeutically homogeneous group than the other tests, which is reflected in its claimed indications: patients with estrogen receptor–positive, lymph node–negative, stage I or II disease who receive tamoxifen. MammaPrint has been tested in diverse populations including a mix of treated and untreated patients, patients with lymph node–negative and –positive disease, and those with estrogen receptor–positive and –negative disease. Accordingly, the claimed indications are broader than those for Oncotype DX. Although the indications for MammaPrint match the populations in whom the test has been evaluated, whether these women should be considered a prognostically homogenous group is a critical question. The H/I test claims to be indicated in patients similar to the Oncotype DX population.
The gene signature underlying H/I has been investigated in large, heterogeneous populations, and differences were found in its prediction ability for specific subgroups. The signature has been variably formulated as a simple ratio or as an index, normalized–different sets of genes or standardized–calibration RNA, and by using stratification thresholds optimized within each study. Whereas plausible mechanisms support the test rationale, the marketed test requires further validation and comparison with conventional combination predictors.
The utility of these tests for clinical decision making raises difficult questions. No study has addressed whether Mammaprint or H/I can predict the clinical benefit of chemotherapy. Oncotype DX is the only gene expression test able to predict clinical benefit of chemotherapy. Although this claim is based on past data, the study design was strong and was based on a randomized, clinical trial of tamoxifen plus chemotherapy versus tamoxifen alone (50). This study suggested that chemotherapy had no benefit in women in the low-risk group, but wide CIs around the observed zero effect does not rule out a meaningful effect.
Even with little data on prediction of treatment benefit, the risk for long-term recurrence or death serves as an effective ceiling on the degree of chemotherapy benefit. If risk is sufficiently low, some patients may forgo chemotherapy. Both the level of the low-risk estimates and the proportion of patients who fall into those categories are therefore of considerable interest. Because various standardized risk prediction tools are freely available, the question is how much these new tests add.
Only a few sources provide evidence on long-term absolute risks after conventional combination risk predictors are taken into account. For Oncotype DX, results of these analyses are published only in abstract form, although the findings are derived from the same NSABP-14 cohort that provided main original validation evidence and are reported by the same authors (46, 47). This showed that Oncotype DX can reclassify patients in highest-risk categories by conventional indices (18% for 2003 St. Gallen, 15% for 2004 National Comprehensive Cancer Network, and 22% for Adjuvant! Online) into clinically relevant lower risks (8%, 8%, and 9%, respectively), although the upper confidence limits on those new lower predictions all exceed 10%. For women at the lowest risk by conventional metrics, being placed in a low-risk stratum seems to lower the risk even further, information that patients might find useful; however, the number of patients in this group and on which this finding is based is small (approximately 30 to 60). For MammaPrint, the only reclassification data reported are those obtained in combination with Adjuvant! Online for a 10-year outcome (36). Adjuvant! Online had no predictive power for survival after the data were stratified by MammaPrint risk group and it had only a very modest effect for 10-year distant recurrence. Similarly, findings were reported for the 70-gene signature in combination with the NIH and St. Gallen criteria (9). The risks in the good-signature MammaPrint groups were higher than the low-risk stratum, in part because the more heterogeneous validation population was at higher risk.
The exact values of the test results provide information that is lost when patients are assigned to risk categories, and the cutoffs for these categories may not correspond to optimal decision thresholds, particularly in combination with other predictors. How the results of such tests are conveyed to and understood by patients and physicians—for example, as absolute probabilities or as qualitative descriptors (low risk)—is critical because these tests are more widely used.
The ideal assessment of value of these tests would be to randomly assign patients to use them or not, as part their therapeutic decision making. The 2 ongoing prospective randomized trials, TAILORx (Trial Assigning IndividuaLized Options for Treatment [Rx]) (56) and MINDACT (Microarray In Node-negative Disease may Avoid ChemoTherapy) (57), do not use such a design. The TAILORx compares disease-free survival among women with previously resected axillary node–negative breast cancer who had an Oncotype DX recurrence score between 11 and 25 and received adjuvant chemotherapy and hormonal therapy versus women who received hormonal therapy alone. All patients receive the test (56); those with a score of 25 or less do not receive chemotherapy, and those with score greater than 25 receive chemotherapy. These thresholds are lower than those conventionally used to designate high risk (score 30) and intermediate risk (score >18 but <30).>Table 3.
As these tests are modified and new ones are marketed, questions will arise about how the tests compare with one another and whether combining the tests has value. Answering these questions will requite comparative effectiveness research. The U.S. government and industry does not traditionally fund comparative effectiveness studies because such studies may not have as much therapeutic promise as new discoveries and because industry is not eager to fund direct comparisons with competitive products. This same dynamic could take hold in the risk-prediction arena. Oversight of test development or research funding should encourage, contrasts with existing expression-based predictors early in the development process. Otherwise, new tests that all claim to offer similar guidance, or perhaps new guidance in previously neglected clinical subsets, will flood the market, and physicians and patients will have no way to evaluate the claims.
In conclusion, the introduction of gene expression tests has ushered in a new era in which many conventional clinical markers may be seen merely as surrogates for more fundamental genetic and physiologic processes that can be measured with these tests. The multidimensional nature of these predictors demands that large numbers of clinically homogeneous patients to be used in the validation process and that exceptional rigor and discipline be applied in evaluation. Every study provides an opportunity to modify a genetic signature, but we must find the right balance between speed of innovation and development of reliable tools. It is important to collect genetic and clinical information from tested patients to facilitate further innovation and thorough evaluation in current patients. Although these tests show great promise to improve predictions of prognosis and treatment benefit for women with early-stage breast cancer, more needs to be learned about the extent of that improvement, in whom they affect, and how they are best incorporated into decision making about current breast cancer treatment.
Glossary
TopMethodsResultsDiscussionGlossaryAuthor & Article InfoReferencesDNA microarray (also called gene chip or DNA chip): A collection of microscopic DNA spots (defined as features), commonly representing single genes or transcripts, arrayed on a solid surface by covalent attachment to chemically suitable matrices, or directly synthesized on them. DNA microarrays use DNA as part of their detection system. Qualitative or quantitative measurements with DNA microarrays use the selective nature of DNA–DNA or DNA–RNA hybridization under high-stringency conditions and fluorophore-based detection. DNA arrays are commonly used for gene expression profiling, i.e., monitoring expression levels of thousands of genes simultaneously, or for comparative genomic hybridization.
Gene expression: The translation of the information encoded in a gene into an RNA transcript. Expressed transcripts include messenger RNAs translated into proteins, as well as other types of RNA, such as transfer RNA, ribosomal RNA, micro RNA, and non-coding RNA, which are not translated into protein. Gene expression is a highly specific process by which cells switch genes on and off in a timely manner, according to their state. The study of messenger RNAs expression in a cell is an indirect way to study the proteins counterpart.
Gene expression profiling: Any genomic technique that measures the fraction of the genes that is expressed in a specific sample. Currently, this definition refers to techniques that allow the assessment of more then 1 gene at a time, especially microarray and real-time reverse transcriptase polymerase chain reaction.
Gene expression profile: Any set of genes for which it is known the expression in a specific sample. A gene expression profile may account for a variable number of genes, and the corresponding expression values may be obtained by different techniques. Gene expression profiles can be associated, by various techniques, to phenotypes.
Gene expression pattern: This is an equivalent definition currently used to refer to gene expression profile.
Gene expression signature: This is an equivalent definition currently uses to refer to a specific gene expression profile, usually associated to a specific phenotype.
Polymerase chain reaction: A molecular biology technique for isolating and exponentially amplifying a DNA sequence of interest in vitro by means of enzymatic replication. This technique has been extensively modified to do a wide array of tasks, and it is now a common tool used in medical and biological research. The polymerase chain reaction is currently used to obtain the sequence of genes to diagnosis hereditary diseases, identify genetic fingerprints (forensics medicine), detect infectious diseases, and create transgenic organisms. Coupled with reverse transcription, it is used to amplify RNA molecules.
Real-time reverse transcriptase polymerase chain reaction: A molecular biology technique that allows the amplification and the quantification in real time of defined RNA molecules from specific specimens. This technology has been used for several years in research and clinical settings to measure RNA molecules. In brief, in the first step, DNA copies of the investigated RNA molecules present in the template are obtained by a reaction named reverse transcription, then DNA amplification is obtained by using polymerase chain reaction, whereas the quantification of the accumulating DNA product is accomplished by the use of specific fluorescent reagents. In this technique the quantification of the target RNA molecule is based on the analysis of the accumulation curve of the complementary DNA, as measured by the fluorescence detected at each cycle of the reaction.
Reverse transcription: In biochemistry, the enzymatic reaction carried on by the RNA-dependent DNA polymerase. This enzyme, also known as reverse transcriptase, is a DNA polymerase enzyme that copies single-stranded RNA into DNA. This process is the reverse of normal transcription, which involves the syntheseis of RNA from DNA.
Transcription: The process by which DNA sequences are copied into complementary RNA molecules by the enzyme RNA polymerase. This reaction represents the transfer of genetic information from DNA into RNA, which is from storing to function. The DNA sequence that is transcribed into an RNA molecule is called a transcript.
Author and Article Information
TopMethodsResultsDiscussionGlossaryAuthor & Article InfoReferencesFrom Johns Hopkins .
Current Author Addresses: Drs. Marchionni and Wolff: Johns Hopkins University, School of Medicine, Oncology Cancer Biology, Baltimore, MD 21287.
Ms. Wilson, Dr. Marinopoulos, and Dr. Bass: Johns Hopkins University, School of Medicine, General Internal Medicine, Baltimore, MD 21287.
Dr. Parmigiani: Johns Hopkins University, School of Medicine, Oncology Informatics, Baltimore, MD 21287
Dr. Goodman: Johns Hopkins University, School of Medicine, Oncology Biostatistics, Baltimore, MD 21287.
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DERECHO A MORIR CON DIGNIDAD

NEW ENG J MED.Volume 357:408-413
July 26, 2007
Number 4

Cancer and the Constitution — Choice at Life's End
George J. Annas, J.D., M.P.H.
J.M. Coetzee's violent, anti-apartheid Age of Iron, a novel the Wall Street Journal termed "a fierce pageant of modern South Africa," is written as a letter by a retired classics professor, Mrs. Curren, to her daughter, who lives in the United States. Mrs. Curren is dying of cancer, and her daughter advises her to come to the United States for treatment. She replies, "I can't afford to die in America. . . . No one can, except Americans."1 Dying of cancer has been considered a "hard death" for at least a century, unproven and even quack remedies have been common, and price has been a secondary consideration. Efforts sponsored by the federal government to find cures for cancer date from the establishment of the National Cancer Institute (NCI) in 1937. Cancer research was intensified after President Richard Nixon's declaration of a "war on cancer" and passage of the National Cancer Act of 1971.2 Most recently, calls for more cancer research have followed the announcement by Elizabeth Edwards, wife of presidential candidate John Edwards, that her cancer is no longer considered curable.
Frustration with the methods and slow progress of mainstream medical research has helped fuel a resistance movement that distrusts both conventional medicine and government and that has called for the recognition of a right for terminally ill patients with cancer to have access to any drugs they want to take. Prominent examples include the popularity of Krebiozen in the 1950s and of laetrile in the 1970s. As an NCI spokesperson put it more than 20 years ago, when thousands of people were calling the NCI hotline pleading for access to interleukin-2, "What the callers are saying is, `Our mother, our brother, our sister is dying at this very moment. We have nothing to lose.'"2 Today, families search the Internet for clinical trials, and even untested chemicals such as dichloroacetate, that seem to offer them some hope. In addition, basing advocacy on their personal experiences with cancer, many families have focused their frustrations on the Food and Drug Administration (FDA), which they see as a government agency denying them access to treatments they need.
In May 2006 these families won an apparent major victory when the Court of Appeals for the District of Columbia, in the case of Abigail Alliance v. Von Eschenbach (hereafter referred to as Abigail Alliance),3 agreed with their argument that patients with cancer have a constitutional right of access to investigational cancer drugs. In reaction, the FDA began the process of rewriting its own regulations to make it easier for terminally ill patients not enrolled in clinical trials to have access to investigational drugs.4 In November 2006, the full bench of the Court of Appeals vacated the May 2006 opinion, and the case was reheard in March 2007.5 The decision of the full bench, expected by the fall, will hinge on the answer to a central question: Do terminally ill adult patients with cancer for whom there are no effective treatments have a constitutional right of access to investigational drugs their physicians think might be beneficial?
The Constitutional Controversy
The Abigail Alliance for Better Access to Developmental Drugs (hereafter called the Abigail Alliance) sued the FDA to prevent it from enforcing its policy of prohibiting the sale of drugs that had not been proved safe and effective to competent adult patients who are terminally ill and have no alternative treatment options. The Abigail Alliance is named after Abigail Burroughs, whose squamous-cell carcinoma of the head and neck was diagnosed when she was only 19 years old. Two years later, in 2001, she died. Before her death she had tried unsuccessfully to obtain investigational drugs on a compassionate use basis from ImClone and AstraZeneca and was accepted for a clinical trial only shortly before her death. Her father founded the Abigail Alliance in her memory.6
The district court dismissed the Abigail Alliance lawsuit. The appeals court, in a two-to-one opinion written by Judge Judith Rogers, who was joined by Judge Douglas Ginsburg, reversed the decision. It concluded that competent, terminally ill adult patients have a constitutional "right to access to potentially life-saving post-Phase I investigational new drugs, upon a doctor's advice, even where that medicine carries risks for the patient," and remanded the case to the district court to determine whether the FDA's current policy violated that right.3
The Right to Life
The appeals court found that the relevant constitutional right was determined by the due-process clause of the Fifth Amendment: "no person shall be . . . deprived of life, liberty, or property without due process of law." In the court's words, the narrow question presented by Abigail Alliance is whether the due-process clause "protects the right of terminally ill patients to make an informed decision that may prolong life, specifically by use of potentially life-saving new drugs that the FDA has yet to approve for commercial marketing but that the FDA has determined, after Phase I clinical human trials, are safe enough for further testing on a substantial number of human beings."3
The court answered yes, finding that this right has deep legal roots in the right to self-defense, and that "Barring a terminally ill patient from the use of a potentially life-saving treatment impinges on this right of self-preservation."3 In a footnote, the court restated this proposition: "The fundamental right to take action, even risky action, free from government interference, in order to save one's own life undergirds the court's decision."3 The court relied primarily on the Cruzan case,7 in which the Supreme Court recognized the right of a competent adult to refuse life-sustaining treatment, including a feeding tube:
The logical corollary is that an individual must also be free to decide for herself whether to assume any known or unknown risks of taking a medication that might prolong her life. Like the right claimed in Cruzan, the right claimed by the [Abigail] Alliance to be free of FDA imposition does not involve treatment by the government or a government subsidy. Rather, much as the guardians of the comatose [sic] patient in Cruzan did, the Alliance seeks to have the government step aside by changing its policy so the individual right of self-determination is not violated.3
The appeals court concluded that the Supreme Court's 1979 unanimous decision on laetrile,8 in which the Court concluded that Congress had made no exceptions in the FDA law for terminally ill cancer patients, was not relevant because laetrile had never been studied in a phase 1 trial and because the Court did not address the question of whether terminally ill cancer patients have a constitutional right to take whatever drugs their physicians prescribe.
The Dissent
Judge Thomas Griffith, the dissenting judge, argued that the suggested constitutional right simply does not exist. He noted, for example, that the self-defense cases relied on are examples of "abstract concepts of personal autonomy," and cannot be used to craft new rights. As to the nation's history and traditions, he concluded that the FDA's drug-regulatory efforts have been reasonable responses "to new risks as they are presented."3 Accepting his argument leaves the majority resting squarely on Cruzan and the laetrile case. As to Cruzan, the dissent argued that "A tradition of protecting individual freedom from life-saving, but forced, medical treatment does not evidence a constitutional tradition of providing affirmative access to a potentially harmful, even fatal, commercial good."3 As to the laetrile case, the judge noted simply that the Court had agreed with the FDA that, "For the terminally ill, as for anyone else, a drug is unsafe if its potential for inflicting death or physical injury is not offset by the possibility of therapeutic benefit."3,8
Finally, the dissenting judge argued that if the new constitutional right were accepted, it was too vague to be applied only to terminally ill patients seeking drugs that had been tested in phase 1 trials. Specifically, the judge asked, must the right also apply to patients with "serious medical conditions," to patients who "cannot afford potentially life-saving treatment," or to patients whose physicians believe "marijuana for medicinal purposes . . . is potentially life saving?"3 In other words, there is no principled reason to restrict the constitutional right the majority created to either terminally ill patients or to post–phase 1 drugs.
Discussion
The facts as illustrated by stories of patients dying of cancer while trying unsuccessfully to enroll in clinical trials are compelling, and our current system of ad hoc exceptions is deeply flawed. The central constitutional issue, however, rests primarily on determining whether this case is or is not like the right-to-refuse-treatment case of Nancy Cruzan, a woman in a permanent vegetative state whose family wanted tube feeding discontinued because they believed that discontinuation was what she would have wanted. I do not think Abigail Alliance is like Cruzan. Rather, it is substantially identical to cases involving physician-assisted suicide, in which a terminally ill patient claims a constitutional right of access to physician-prescribed drugs to commit suicide.
The Supreme Court has decided, unanimously, that no right to physician-prescribed drugs for suicide exists.9,10 There is no historical tradition of support for this right. And although the right seems to be narrowly defined, it is unclear to whom it should apply — why only to terminally ill patients? Don't patients in chronic pain have even a stronger interest in suicide? Why is the physician necessary, and why are physician-prescribed drugs the only acceptable method of suicide? None of these questions can be answered by examining the Constitution.11
Similarly, in Abigail Alliance, the new constitutional right proposed has no tradition in the United States, and it cannot be narrowly applied. For example, why should a constitutional right apply only to people who have a particular medical status? And why should a physician be involved at all? If patients have a right to autonomy, why isn't the requirement of a government-licensed physician's recommendation at least as burdensome as the requirement of the FDA's approval of the investigational drug? And why would the Constitution apply only to investigational drugs for which phase 1 trials have been completed? Why not include access to investigational medical devices, like the artificial heart, or even to Schedule I controlled substances, like marijuana or lysergic acid diethylamide (LSD)? If it is a constitutional right, these should be available too, at least unless the state can demonstrate a "compelling interest" in regulating them.
My prediction is that after rehearing this case en banc, the full Circuit Court will reject the position of the Abigail Alliance for the same reasons that the Supreme Court rejected the "right" of terminally ill patients to have access to physician-prescribed drugs they could use to end their lives.9,10,11 To decide otherwise would entirely undermine the legitimacy of the FDA. Patients in the United States have always had a right to refuse any medical treatment, but we have never had a right to demand mistreatment, inappropriate treatment, or even investigational or experimental interventions. This will not, however, be the end of the matter. After the physician-assisted–suicide cases, the fight appropriately shifted to the states, although so far only one, Oregon, has provided its physicians with immunity for prescribing life-ending drugs to their competent, terminally ill patients.12 In the Abigail Alliance case, the debate will continue in the forum in which it began — the FDA — and in Congress.
Congress
Congressional action also had its birth with the story of one patient with cancer and was also heavily influenced by another individual patient involved in a controversy over removal of a feeding tube. "Terri's Law" was enacted in Florida in 2003 to try to prevent the removal of a feeding tube from Terri Schiavo; the case was substantially similar to Cruzan. Terri's case gained national attention 2 years later.13 In the midst of it, in March 2005, the Wall Street Journal asserted, in an editorial titled "How About a `Kianna's Law'?," "If Terri Schiavo deserves emergency federal intervention to save her life, people like Kianna Karnes deserve it even more."14 At the time, Kianna Karnes was a 44-year-old mother of four who was dying of kidney cancer. Her only hope of survival, according to the editorial, was to gain access to one of two experimental drugs in clinical trials, but neither of the two companies running the trials (Bayer and Pfizer) would make the drugs available to her on a compassionate-use basis. This was because, according to the Wall Street Journal, the FDA "makes it all but impossible" for the manufacturers "to provide [drugs] to terminal patients on a `compassionate use' basis."14
Almost immediately after the editorial was published, both drug manufacturers contacted Kianna's physicians to discuss releasing the drugs to her. But within 2 days after publication, she was dead. The Wall Street Journal editorialized, "Isn't it a national scandal that cancer sufferers should have to be written about in the Wall Street Journal to be offered legal access to emerging therapies once they've run out of other options?"15 It noted that Mrs. Karnes' father, John Rowe — himself a survivor of leukemia — was working with the Abigail Alliance on a "Kianna's Law." That law, formally titled the "Access, Compassion, Care, and Ethics for Seriously Ill Patients Act" or the "ACCESS Act," was introduced in November 2005 and is an attempt to make it much easier for seriously ill patients to gain access to experimental drugs.16,17
The act begins with a series of congressional findings, including that "Seriously ill patients have a right to access available investigational drugs, biological products, and devices." The act permits the sponsor to apply for approval to make an investigational drug, biologic product, or device available on the basis of data from a completed phase 1 trial, "preliminary evidence that the product may be effective against a serious or life-threatening condition or disease," and an assurance that the clinical trial will continue.17 The patient, who must have exhausted all approved treatments, must provide written informed consent and must also sign "a written waiver of the right to sue the manufacturer or sponsor of the drug, biological product, or device, or the physicians who prescribed the product or the institution where it was administered, for an adverse event caused by the product, which shall be binding in every State and Federal court."17
Although Congress is the proper forum to address this issue, this initial attempt has some of the same problems as the Abigail Alliance decision: the patients to whom it applies are ambiguously classified, and clinical research seems to be equated with clinical care. Also troubling is that the patients (and would-be subjects) are asked to assume all of the risks of the uncontrolled experiments, and current rules of research — which protect subjects by prohibiting mandatory waivers of rights — are jettisoned, with the requirement of such waivers becoming the price of obtaining the investigational agent from an otherwise reluctant drug company.
FDA Proposal
In direct response to Abigail Alliance, the FDA proposed amending its rules to encourage more drug companies to offer their investigational drugs through compassionate-use programs.4 These programs first came into prominence during the early days of infection with the human immunodeficiency virus (HIV) and AIDS, when there were no effective treatments and AIDS activists insisted that they have early access to investigational drugs because, in the words of their inaccurate slogan, "A Research Trial Is Treatment Too."18 Because the FDA could not stand the political pressure generated by the activists, the compassionate-use program was developed as a kind of political safety valve to provide enough exceptions to save their basic research rules. In early December 2006, the FDA continued this political-safety-valve approach by issuing new proposed regulations with a title that could have been taken directly from the AIDS Coalition to Unleash Power (ACT-UP): "Expanded Access to Investigational Drugs for Treatment Use."19
The FDA's expanded-access proposal applies to "seriously ill patients when there is no comparable or satisfactory alternative therapy to diagnose, monitor, or treat the patient's disease or condition."4 Manufacturers are required to file an "expanded access submission," and the product must be administered or dispensed by a licensed physician who will be considered an "investigator," with all the reporting requirements that role entails.3
Whether or not the proposal is adopted, it will do little to increase access, since the major bottleneck in the compassionate-use program has never been the FDA. The manufacturers have no incentives to make their investigational products available outside clinical trials. This is because direct access to investigational drugs by individuals may make it more difficult to recruit research subjects, and thus to conduct the clinical trials necessary for drug approval, and could also subject the drug manufacturer to liability for serious adverse reactions. Even without a lawsuit, a serious reaction to a drug outside a trial could adversely affect the trial itself.4,16,20 The drug companies are right to worry that the approaches of the judiciary, Congress, and the FDA will probably make clinical trials more difficult to conduct, because few seriously ill patients who have exhausted conventional treatments would rather be randomly assigned to an investigational drug than have a guarantee that they will receive the investigational drug their physician recommends for them. This could result in significant delays in the approval and overall availability of drugs that demonstrate effectiveness — a result no one favors. Even if patients with cancer are willing buyers, drug manufacturers are not willing sellers.
Physicians and Patients
The cover story for all the proposed changes is patients' choice. But without scientific evidence of the risks and benefits of a drug, choice cannot be informed, and for seriously ill patients, fear of death will predictably overcome fear of unknown risks. This is understandable. As psychiatrist Jay Katz, the leading scholar on informed consent, has noted, when medical science seems impotent to fight nature, "all kinds of senseless interventions are tried in an unconscious effort to cure the incurable magically through a `wonder drug,' a novel surgical procedure, or a penetrating psychological interpretation."21 Another Wall Street Journal article, entitled "Saying No to Penelope,"22 illustrates the impossibility of limiting access to unproven cancer drugs to competent adults. The article tells the story of 4-year-old Penelope, who is dying from neuroblastoma that has proved resistant to all conventional treatments. Her parents seek "anything [that] has a prayer of saving her." In her father's words, "The chance of anything bringing her back from the abyss now is very low. But the only thing I know for sure is if we don't treat her, she will die." With Penelope hospitalized and in pain, her parents continue "searching Penelope's big brown eyes for clues as to how long she wants to continue to battle for life."
It is suggested that the requirement of a physician's recommendation can safeguard against "magical thinking" and help make informed consent real.23 But as Katz has noted, although physicians (and, he could have added, drug companies) often justify such last-ditch interventions as simply being responsive to patient needs, the interventions "may turn out to be a projection of their own needs onto patients."21
Government and the Market
Another recurrent theme is the belief that government regulation is evil, a central tenet of the laetrile litigation of the 1970s. The court hearing Abigail Alliance was correct to note that laetrile never underwent a phase 1 trial, but every indication was that the drug, also known as vitamin B17, was harmless, albeit also ineffective against cancer. Laetrile became a legal cause celebre in 1972, when California physician John A. Richardson was prosecuted for promoting laetrile. Richardson was a member of the John Birch Society, which quickly formed the Committee for Freedom of Choice in Cancer Therapy, with more than 100 committees nationwide.24 It took another 7 years before the FDA prevailed in its case against laetrile before the Supreme Court.8 The basic arguments against FDA regulation remain the same today: the FDA follows a "paternalistic public policy that prevents individuals from exercising their own judgment about risks and benefits. If the FDA must err, it should be on the side of patients' freedom to choose."25
Public Policy
The FDA will prevail again today, not only because there is no constitutional right of access to unapproved drugs but also because even if there were, the state has the same compelling interest in approving drugs as it has in licensing physicians. From a public policy view, the Abigail Alliance court, the Congress, and the FDA all seem to be suffering from the "therapeutic illusion" in which research, designed to test a hypothesis for society, is confused with treatment, administered in the best interests of individual patients.21,26,27 Of course there is a continuum, and it is perfectly understandable that many patients with cancer, told that there is nothing conventional medicine can do for them, will want access to whatever is available in or outside the context of clinical trials. But this is a problem for patients, physicians, the FDA, and drug manufacturers. First, because terminally ill patients can be harmed and exploited, there are better and worse ways to die.21,26 Second, it is only through research, not "treatment," that cancer may become a chronic illness that is treated with a complex array of drugs, given either together or in a progression.28,29 The right to choose in medicine is a central right of patients, but the choices can and should be limited to reasonable medical alternatives, which themselves are based on evidence.
This is, I believe, good public policy. But it is also much easier said than done.30 Death is feared and even dreaded in our culture, and few Americans are able to die at home, at peace, with our loved ones in attendance, without seeking the "latest new treatment." There always seems to be something new to try, and there is almost always anecdotal evidence that it could help. This is one reason that even extremely high prices do not affect demand for cancer drugs, even ones that add little or no survival time.31,32 When does caring for the patient demand primary attention to palliation rather than to long-shot, high-risk, investigational interventions? Coetzee's Mrs. Curren, who rejected new medical treatment for her cancer and insisted on dying at home, told her physician, whom she saw as "withdrawing" from her after giving her a terminal prognosis — "His allegiance to the living, not the dying" — "I have no illusions about my condition, doctor. It is not [experimental] care I need, just help with the pain."1
From the Department of Health Law, Bioethics, and Human Rights, Boston University School of Public Health, Boston.
References
Coetzee JM. Age of iron. London: Seeker & Warburg, 1990.
Patterson JT. The dread disease: cancer and modern American culture. Cambridge, MA: Harvard University Press, 1987.
Abigail Alliance v. Von Eschenbach, 445 F.3d 470 (DC Cir 2006). Vacated 469 F.3d 129 (DC Cir 2006).
Proposed rules for charging for investigational drugs and expanded access to investigational drugs for treatment use. Rockville, MD: Food and Drug Administration, 2006. (Accessed July 6, 2007, at http://www.fda.gov/cder/regulatory/applications/IND_PR.htm.)
Abigail Alliance v. Von Eschenbach, 429 F.3d 129 (DC Cir 2006).
Jacobson PD, Parmet WE. A new era of unapproved drugs: the case of Abigail Alliance v Von Eschenbach. JAMA 2007;297:205-208. [Free Full Text]
Cruzan v. Director, Missouri Dept. of Health, 497 U.S. 261 (1990).
United States v. Rutherford, 442 U.S. 544 (1979).
Washington v. Glucksberg, 521 U.S. 702 (1997).
Vacco v. Quill, 521 U.S. 793 (1997).
Annas GJ. The bell tolls for a constitutional right to assisted suicide. N Engl J Med 1997;337:1098-1103. [Free Full Text]
Brugarolas J. RenPhase 1, 2 , 3 and 4 trials
This page is about the different phases of clinical trials. It has information on
Phase 1 trials
Phase 2 trials
Phase 3 trials
Phase 4 trials
Phase 1 (phase I)These are the earliest trials in the life of a new drug or treatment. They are usually small trials, recruiting anything up to about 30 patients, although often a lot less. The trial may be open to people with any type of cancer. When laboratory testing shows a new treatment might help treat cancer, phase 1 trials are done to find out
The safe dose range
The side effects
How the body copes with the drug
If the treatment shrinks cancer Patients are recruited very slowly onto phase 1 trials. So although they don't recruit many patients, they can take a long time to complete. The first few patients to take part (called a 'cohort' or group) will be given a very small dose of the drug. If all goes well, the next group will get a slightly higher dose. The dose will gradually be increased with each group. The researchers will monitor the effect, until they find the best dose to give. This is called a 'dose escallation study'. In a phase 1 trial, you may have lots of blood tests, as the researchers look at how the drug is affecting you. And at how your body copes with, and gets rid of the drug. They will also record any side effects.People entering phase 1 trials often have advanced cancer and have usually had all the treatment available to them. This is because they may benefit from the new treatment in the trial, but many won't. The aim of the trial is to look at doses and side effects. This work has to be done first, before we can test the potential new treatment to see if it works. Phase 1 trials are important because they are the first step in finding new treatments for the future.
Phase 2 trials (phase II)Not all treatments tested in a phase 1 trial make it to a phase 2 trial. These trials may be done on people who all have same type of cancer, or with several different types of cancer. Phase 2 trials are done to find out
If the new treatment works well enough to test in a larger phase 3 trial
Which types of cancer the treatment works for
More about side effects and how to manage them
More about the best dose to use Although these treatments have been tested at phase 1, you may still have side effects that the doctors don't know about. Drugs can affect people in different ways. Phase 2 trials are often larger than phase 1. There may be up to 50 or so people taking part. If the results of phase 2 trials show that a new treatment may be as good as existing treatment, or better, it then moves to phase 3.
Phase 3 (phase III)These trials compare new treatments with the best currently available treatment (the standard treatment). They may compare
A completely new treatment with the standard treatment
Different doses or ways of giving a standard treatment
A new radiotherapy schedule with the standard onePhase 3 trials are usually much larger than phase 1 or 2. This is because differences in success rates may be small. So, you would need many patients in the trial to show the difference.For example, 6 out of 100 (6%) more people get a remission with a new treatment compared to standard treatment. If there were 50 people in the new treatment group and 50 people in the standard treatment group, there may be 3 more people in remission in the new treatment group. The 2 groups would not look that different. But if they gave each treatment to 5,000 people, there could be 300 more remissions in the new treatment group.Sometimes phase 3 trials involve thousands of patients in many different hospitals and even different countries.
RandomisationPhase 3 trials are usually randomised. This means the researchers put the people taking part into 2 groups at random. One group gets the new treatment and the other the standard treatment. There is more about randomisation and different types of trials in this section.
OverviewsTrial overviews are studies that combine all the results from phase 3 trials of a new treatment. They are sometimes called meta-analyses. The idea is to get a broader picture of how well a treatment works. The more data (information) you have, the more accurate the results are likely to be.
Phase 4 (phase IV)Phase 4 trials are done after a drug has been shown to work and has been granted a license. So they are looking at drugs that are already available for doctors to prescribe, rather than new drugs that are still being developed.The main reasons pharmaceutical companies run phase 4 trials are to find out
More about the side effects and safety of the drug
What the long term risks and benefits are
How well the drug works when it’s used more widely than in clinical trialsThere is more information about the different phases of clinical trials on the website of The Association of the British Pharmaceutical Industry (ABPI).

al-cell carcinoma -- molecular pathways and therapies. N Engl J Med 2007;356:185-186. [Free Full Text]
Callahan D. False hopes: why America's quest for perfect health is a recipe for failure. New York: Simon and Schuster, 1998.
Berenson A. Hope, at $4,200 a dose: why a cancer drug's cost doesn't hurt demand. New York Times. October 1, 2006:BU1.
Anand G. From Wall Street, a warning about cancer drug prices. Wall Street Journal. March 15, 2007:A1.

TRASTORNOS DE LA EYACULACION

Ejaculatory disorders: pathophysiology and management
Carlo Bettocchi, Paolo Verze, Fabrizio Palumbo, Davide Arcaniolo and Vincenzo Mirone*
Correspondence *University of Naples Federico II, Department of Urology, Via Sergio Pansini 5, 80131 Naples, Italy Email mirone@unina.it

Summary
Ejaculatory dysfunction (EjD) is one of the most common male sexual disorders, yet EjD is still frequently misdiagnosed or overlooked as a result of numerous patient and physician barriers. The wide spectrum of EjD ranges from premature or rapid ejaculation, through delayed ejaculation, to a complete inability to ejaculate—otherwise known as anejaculation—and includes retrograde ejaculation and painful ejaculation. Conventional algorithms for managing ejaculatory disorders are based either on an organic or psychogenic etiology, with the latter more traditionally considered the main cause. This paper reviews physiopathological, diagnostic and therapeutic aspects of ejaculation disorders, with a particular focus on the most prevalent disorder, premature ejaculation.
Keywords:
anejaculation, ejaculatory dysfunction, intravaginal ejaculatory latency time, premature ejaculation, selective serotonin reuptake inhibitors
Introduction
As has been previously established, a normal sexual response cycle comprises four interactive, nonlinear stages: desire, arousal, orgasm and resolution. In males, orgasm usually coincides with ejaculation, but represents a distinct cognitive and emotional cortical event. Ejaculatory dysfunction (EjD) is one of the most common male sexual disorders. Fertility is a major concern for younger men, while EjD can cause considerable distress to men of all ages. In a recent survey of 12,815 men aged 50–80 years, 46% reported an ejaculatory disturbance within the previous 4 weeks and 59% were particularly bothered by it, especially when occurring concomitantly with lower urinary tract symptoms.1
The wide spectrum of EjD ranges from premature or rapid ejaculation, through delayed ejaculation, to a complete inability to ejaculate—known as anejaculation—and includes retrograde ejaculation and painful ejaculation.2
Delayed ejaculation (DE), or ejaculatory insufficiency, is defined as the inhibition of the ejaculatory reflex, with absent or reduced seminal emission and impaired ejaculatory contractions, possibly occurring concomitantly with reduced or absent orgasm. Delayed ejaculation affects approximately 4% of sexually active men.3 Retrograde ejaculation, the most medically concerning disorder of sperm emission, occurs in 75% of men who have undergone transurethral resection of the prostate and, to a lesser extent, following bladder neck incision, owing to iatrogenic bladder neck incompetence in both cases.4
The most common EjD is premature ejaculation (PE).5, 6 According to the authors of a comprehensive review of the literature, PE affects 5–40% of sexually active men.7 This disorder is most frequently reported by adolescents or young adults, and affects more men from East Asia and fewer men from Middle Eastern and African countries than in other regions. The European prevalence seems to lie between that of East Asia and Middle Eastern and African countries.6
Still, epidemiological data on PE have been difficult to accrue owing to the lack of a globally accepted definition of the disorder.5 Criteria have been published that define any ejaculation occurring in 1 min, 2 min, 3 min or even 7 min from penetration, or 8–15 penile thrusts, as premature.6 Alternatively, the European Association of Urology disorders of ejaculation guidelines, published in 2004, define PE as the inability to control ejaculation for a "sufficient" length of time before vaginal penetration.8 Although there is no universally accepted meaning of a "sufficient" length of time, this should include those patients who are not able to delay ejaculation for more than a few coital thrusts, or even before vaginal penetration.8 Masters and Johnson9 have suggested that a man has PE if he is unable to delay his ejaculation until his partner is sexually satisfied in at least 50% of their sexual approaches.
The absence of a consensus medical definition for PE encourages a "patient-dependent" definition and a "patient-decided" diagnosis.6 This approach is risky, because diagnosis and possible therapy would then be based solely on subjective parameters, which are clearly influenced by culture, religion, policy, society, and the media—all aspects that greatly deviate from a medical definition.6 Hence, in 1994, Waldinger et al.10 proposed a simple objective method to define PE termed the intravaginal ejaculation latency time (IELT), which is the time from the start of vaginal intromission to the start of intravaginal ejaculation. For clinical assessment and therapeutic monitoring, this method could be considered the most objective in evidence-based sexual medicine.11
In 2000, the Diagnostic and Statistical Manual of Mental Disorders, version 4 (DSM-IV) expanded the definition of PE to be persistent or recurrent ejaculation with minimal sexual stimulation before, upon, or shortly after penetration and before the person wishes it, and noted an association with marked distress or interpersonal difficulty.12 In accordance with this definition, PE presents with three components: a short time interval between penetration and ejaculation, the inability to control ejaculation and distress for the man or for the couple.
Physiology of ejaculation
The ejaculatory process is mainly mediated by the autonomic nervous system,13, 14 and consists of two main phases: emission and expulsion.15 Two groups of anatomical structures are specifically involved and distinguished in each phase. The organs involved in the emission phase comprise the epididymis, vas deferens, seminal vesicles, prostate gland, prostatic urethra and bladder neck. The organs participating in the expulsion phase include the bladder neck and urethra, as well as the pelvic striated muscles.15
Emission phase
During the emission phase, spermatozoa mixed with products secreted by accessory sexual glands are ejected into the posterior urethra by sequential epithelial secretion and smooth muscle cell contractions. Organs involved in this phase have a dense sympathetic and parasympathetic innervation mainly deriving from the pelvic plexus, also referred to as the inferior hypogastric plexus in humans. The nerves are situated retroperitoneally within the sagittal plane on either side of the rectum, and lie lateral and posterior to the seminal vesicles.15 The principle neurotransmitter involved in the sympathetic stimulation is norepinephrine, and is balanced by parasympathetic mediators such as acetylcholine. Results from rat studies have suggested a role for oxytocin and nonadrenergic/noncholinergic (NANC) factors, including ATP, neuropeptide Y (NPY), vasoactive intestinal peptide (VIP) and nitric oxide (NO) in ejaculation.16, 17, 18, 19 Sympathetic fibers, originating from the D10–L2 medullar center, are crucial to maintaining physiologic ejaculatory function. These nerves originate from the lumbar ganglia of the paravertebral sympathetic trunk. They pass posteriorly to the vena cava and then into the interaortocaval space (on the right side) or laterally to the aorta (on the left side). They are the principal constituents of the superior hypogastric plexus. Many surgical operations can cause an ejaculation disorder by disturbing these neuronal connections; importantly, retroperitoneal lymphadenectomy for testis cancer tends to affect young males, and has been the subject of several studies.20 The anatomosurgical concepts used to spare ejaculation function during retroperitoneal lymphadenectomy can be adopted for other retroperitoneal surgical operations that might also result in ejaculation disorders, such as wide lymphadenectomy for renal cell carcinoma or tumors of the upper urinary tract, exeresis of preaortic tumors, exeresis or disjunction of horseshoe kidney, and aortoiliac revascularization.20
Expulsion phase
During the expulsion phase, sperm is ejected from the urethra through the glans meatus. According to the commonly accepted theory, expulsion is a spinal cord reflex that occurs as the ejaculatory process reaches a 'point of no return'. During expulsion, the smooth muscle fibers of the bladder neck contract to prevent the backward flow of semen into the bladder; the pelvic floor muscles, along with the bulbospongiosus and ischiocavernosus muscles, have primary roles in this function, and display stereotypical rhythmic contractions to propel semen distally throughout the bulbar and penile urethra. During this process, the external urinary sphincter is normally relaxed.21 The bladder neck and proximal portion of the urethra, which both contain abundant smooth muscle fibers, receive dual sympathetic and parasympathetic innervation. Besides the cholinergic and noradrenergic components, the principal mediators found in these nerve terminals are NPY, VIP, and NO. The external urethral sphincter and pelvic floor striated muscles are solely regulated by the somatic nervous system, but there is no definitive evidence that a voluntary control of the expulsion phase exists in humans. Rhythmic pelvic striated muscle contractions leading to sperm expulsion are probably triggered mainly by the presence of semen in the bulbous urethra,22 although several alternative theories have been formulated and could explain the preservation of rhythmic contractions in patients who have dry ejaculations or patients who have undergone radical prostatectomy for prostate cancer.
In fact, while these stimuli might be sufficient to elicit ejaculation in experimental conditions, experimental and clinical findings do not support seminal fluid build-up as a necessary trigger.23 In a rat model, anesthetizing the urethra or decreasing seminal emission by injection of guanethidine monosulfate did not prevent the ejaculatory pattern during copulation.24 Moreover, ejaculatory motor patterns were still observed in patients after cystectomy, prostatectomy, vesiculectomy and urethrectomy.25 Likewise, ejaculatory motor patterns were not affected in male rats by removal of the seminal vesicles26 or all accessory genital glands.27 Thus, it might be possible that seminal emission contributes to the sensory stimuli that trigger ejaculation in intact, normal males, but it is not a necessary sensory stimulus. Many questions still remain unanswered as to the exact sensory mechanisms responsible for erection, emission and expulsion.23
Both experimental and clinical evidence suggests the presence of a spinal ejaculatory generator, which integrates peripheral and central stimuli and exerts both somatic and parasympathetic efferent pathways. The integrity of these spinal nuclei is necessary and sufficient for ejaculation, as demonstrated by the induction of the ejaculatory process after peripheral stimulation in animals and humans with spinal cord injuries located above these nuclei. The ability of peripheral stimuli to induce a complete human or partial rat penile ejaculation, despite the loss of reciprocal connections with supraspinal structures, implies that the spinal cord contains the complete neural machinery necessary for ejaculation.28 Studies using the expression of the protein product of the immediate-early gene c-fos as a marker of neural activation have demonstrated that some neurons in the central gray matter of the lumbar spinal cord are specifically activated following mating.29 A major breakthrough came from the identification of a population of neurons that have a pivotal role in the generation of an ejaculation.30 This population of neurons consists of cells located in lamina X and the medial portion of lamina VII of lumbar segments 3 and 4 (L3–4).30 Based on their location and thalamic projections, these particular neurons are referred to as lumbar spinothalamic cells.28
Furthermore, the spinal ejaculation generator is under the inhibitory and excitatory influences of supraspinal sites, including the nucleus paragigantocellularis, the paraventricular nucleus of the hypothalamus and the medial preoptic area.23 A modulating role for supraspinal areas in the ejaculatory threshold was confirmed by the finding that the urethrogenital reflex cannot be elicited in intact rats, but usually requires either thoracic spinal transection or a lesion within the nucleus paragigantocellularis, an area in the ventrolateral medulla in the brainstem.31 Definitive information on areas in the human brain that lead to ejaculation is sparse, although several studies using PET promise to provide further knowledge soon.
Serotoninergic control of ejaculation
Ejaculation involves cerebral sensory areas, motor centers and several spinal nuclei that are tightly interconnected, underlying a complex neurochemical modulation of the process. Amongst a wide number of neurotransmitters, including serotonin (5-HT), dopamine, oxytocin, -aminobutyric acid (GABA), adrenaline, acetylcholine and NO, which have been shown to be involved in the regulation of ejaculation, 5-HT has a primary role at several levels of the neuraxis.15, 32
According to the principle outcomes of several experimental and clinical models, the overall effect of 5-HT on ejaculation is inhibitory.32, 33 Marson and McKenna31, 34 have shown that intrathecal injection of 5-HT inhibits ejaculation, while other analyses have confirmed the prolongation of ejaculation latency after central serotonin injection.35, 36 Yet, the local injection of low doses of 5-HT into the dorsal or median raphe nuclei lowers the ejaculatory threshold, which facilitates male rat ejaculatory behaviors.37 Theoretically, this particular finding is due to activated feedback systems that inhibit cell firing and decrease 5-HT levels in neuronal projections.38
Different 5-HT receptor subtypes might have an opposite action on the central command of ejaculation. Amongst the 14 structurally distinct 5-HT receptor subtypes identified to date, 5-HT1A, 5-HT1B and 5-HT2C subtypes, and perhaps 5-HT7 subtypes, have been shown to be involved in the control of the ejaculatory response. Animal studies have shown that administration of 5-HT1A-receptor agonists exerts an overall facilitating effect on ejaculation. Conversely, it has also been shown in several studies that subcutaneous administration of 5-HT1B receptor agonists (e.g. anpirtoline and TFMPP) impairs ejaculation in rats.39
Pathophysiology of premature ejaculation
Studies have been performed to investigate the etiology and pathogenesis of PE, but clear mechanisms have yet to be defined. The pathophysiology of PE can be differentiated into five distinct areas—psychorelational, neurobiological, urological, hormonal and andrological—which, consequently, suggests five different therapeutic approaches.40
Pathogenesis from dysfunctional psychorelational concepts is the most studied cause of PE. According to this theory, distortions of beliefs and false convictions about sexuality, established in childhood as a consequence of adverse influences on sexual behavior, might lead to sexual dysfunction such as PE. During early sexual experiences PE is frequent and might even be considered normal. In men with PE, however, they might not allow themselves to receive the sensory feedback of those sensations occurring immediately before orgasm, which would enable the ejaculatory reflex to be brought under voluntary control.41 Also, anxiety (general or related to sexual performance), guilt (from a belief that sexual activity is sinful), and fear (of pregnancy, sexually transmitted diseases and/or being discovered) have frequently been suggested as causes of PE.42
Contrary to the classic psychosexual approach, Waldinger et al.43 have theorized, on the basis of animal models, that PE is not a psychological disorder but rather a neurobiological phenomenon due to chronic (genetic or acquired) central serotoninergic hypoactivity. Nevertheless, animal studies cannot be easily extrapolated to human sexual behavior, and further research is needed to clarify the role of the serotoninergic system in the pathogenesis of PE. Among peripheral neurobiological causes, major neurological disorders—such as multiple sclerosis, spina bifida and spinal cord tumors—are rare. Clinical studies have demonstrated that a short frenulum, penile hypersensitivity and reflex hyperexcitability could be important contributing factors to PE.44, 45 Nevertheless, Vanden Broucke et al.46 has demonstrated that penile sensitivity measurements do not correlate with ejaculation latency times (ELT) in men with normal sexual function; therefore, ELT variability cannot be explained by differences in penile sensitivity, at least in normal men. Moreover, the reproducible variation in ELT probably cannot be explained by differences in peripheral sensation, although this does not exclude the possibility that penile sensitivity has a role in creating a significantly lowered ejaculatory threshold in patients with PE.46
Sometimes urological or andrological disease can lead to PE. There is a high prevalence of chronic prostatitis in men with PE (56.5%), suggesting that prostatic inflammation is a predisposing condition for PE.47 In many cases PE is the only complaint presented, but patients frequently present with other sexual problems, such as erectile dysfunction (ED). It could be inferred that PE and ED share a vicious cycle, in which a man trying to control his ejaculation instinctively reduces his level of excitation (which can lead to ED), and a man trying to achieve an erection attempts to increase his excitation (which can lead to PE). In addition, many patients with ED might ejaculate early to hide the weakness of their erection.40
Recently, hyperthyroidism has been shown to be a possible promoting factor of PE.48 As the relationship between thyroid hormones and ejaculatory dysfunction is currently unknown, three possible sites of action have been hypothesized: the sympathetic nervous system, the serotoninergic system and the endocrine/paracrine system.40 In addition, thyroid hormone receptors have been identified in animal and human testes, and might also be present in other male genital tract structures that trigger ejaculation.49
Diagnostic aspects of ejaculatory disorders
Premature ejaculation
Diagnosis of PE in clinical practice is straightforward, as it is simply based on patient self-report, clinical history, sexual history and examination findings; however, primary and acquired PE must be differentiated, which are respectively defined as experience with the problem for as long as the man has been sexually active, and development of the condition after having had previous satisfying sexual relationships without problems.
Obtaining a clinical history is crucial for identifying potential comorbidities, such as diabetes, neuropathies, traumas or urogenital infections, as well as previous surgery or drug consumption. Particular attention should be paid to micturition and ejaculation characteristics (such as nocturnal emissions, ejaculatory ability in particular circumstances, congenital or acquired disorders, and the evolution of dysfunction), as well as to the psychosexual atmosphere (education, features of the affective relationship, pre-existing psychological trauma, and previous psychological therapy).8 Owing to the absence of validated tools for diagnosing PE, the American Urological Association guidelines recommend that the diagnosis of PE be based solely upon information gathered from taking a sexual history, such as questions concerning the frequency and duration of PE, the proportion of sexual attempts with PE, the patient's relationships with specific partners, the frequency and nature of sexual activity, and the effects of PE on sexual activity and quality of life.50
Genital and rectal physical examinations are necessary to establish the absence of penile shaft abnormalities (i.e. short frenulum) or prostatic inflammation; the latter requires the performance of transrectal ultrasonography and a Meares–Stamey test. Laboratory tests are required only when there is suspicion of illicit drug use or excessive alcohol consumption.
The major objective when diagnosing PE is to quantify the length of time between penetration and ejaculation, although a multidimensional assessment of patients affected with PE, including psychosocial involvement, is also needed. Various subjective, self-report questionnaires have been proposed, and many are universally accepted and used in clinical practice. One of the most widely-used questionnaires is the Premature Ejaculation Diagnostic Tool (PEDT), a five-item tool developed to systematically apply the Diagnostic and Statistical Manual of Mental Disorders, revised version 4 (DSM-IV-TR) criteria for diagnosing the presence or absence of PE. The historical development of the DSM and the International Classification of Diseases (ICD) categorization of mental disorders have been critically reviewed. The critical reanalysis of two previous studies using the DSM-IV-TR definition of PE revealed a low, positive-predictive value for the DSM-IV-TR definition when used as a diagnostic test.51 By contrast, Symonds et al.52 confirm that the PEDT is an extensively validated self-report measure that can systematically assess DSM-IV-TR criteria to provide accurate diagnoses in patients affected with PE. Another newly developed tool for diagnosing PE has been proposed by Wang and colleagues;53 the Chinese Index of Premature Ejaculation (CIPE) is a 10-item, structured questionnaire that explores the major male sexual cycle—seemingly an ideal basis for the diagnosis of PE. Furthermore, an abridged version of the 25-item Male Sexual Health Questionnaire (MSHQ) was developed and validated (MSHQ-EjD Short Form) for assessing EjD.54 The MSHQ-EjD Short Form contains three ejaculatory function items and one ejaculation bother item; it offers excellent psychometric properties and should be a useful instrument for assessing EjD in clinical settings.
IELT, expressed in seconds or minutes, is the most widely used measurement to assess PE. A recent multinational, community-based, age-ranging study demonstrated that the distribution of the IELT was positively skewed, with a median time of 5.4 min and a range of 0.55–44.1 min. The differences between the men of the five different countries were explored; for all the men considered, the median IELT values were independent of condom usage. In countries excluding Turkey, the median IELT values were also independent of circumcision status.55
IELT is usually measured with a stopwatch operated by the female partner, but owing to the wide influences of psychosocial factors and cultural backgrounds, an objective evaluation is often lacking. A novel apparatus has been designed and validated to collect electronic data on IELT, called the Sexual Assessment Monitor (SAM). Dinsmore et al.56 showed that the SAM is able to measure effectively and safely the time to ejaculation from the start of vibration in healthy volunteers and men with PE. The authors concluded that the SAM has the potential to become the gold standard for the diagnosis of PE, as well as in the design of clinical trials.
Neurophysiologic tests, such as pudendal nerve somatosensory evoked potentials, bulbocavernosus (BC) reflex and BC perineal motor evoked potentials, have been recently assessed for use in the investigation of the somatic sensory and motor function of the genital area in patients presenting with primary PE. Neither an increased speed of conduction along the pudendal sensory pathway nor an increased cortical representation of the sensory stimuli from the genital area—such as hyperexcitability of the BC reflex—were confirmed in successive studies. Despite great initial interest in this area, an electrophysiologic approach by itself does not seem sufficient to identify the cause of primary PE.57
Delayed ejaculation and anejaculation
Although no clear criteria exist, given that most sexually functional men ejaculate within about 3–8 min following intromission, men with latencies beyond 20–30 min and consequent distress or men who simply cease sexual activity due to exhaustion or irritation qualify for a diagnosis of delayed ejaculation.58
The diagnostic evaluation of delayed ejaculation or anejaculation focuses on finding potential physical causes and specific psychological or acquired causes of the disorder. A medical examination and patient history are crucial, as these might uncover physical anomalies, pathophysiologies and/or iatrogenically induced conditions (e.g. treatment with antihypertensive, antipsychotic or antidepressant drugs, surgical procedures such as retroperitoneal lymphadenectomy or aortoiliac or colorectal surgery) associated with delayed or absent ejaculation. Particular attention should be given to identifying reversible urethral, prostatic, epididymal and/or testicular infections. Evaluation typically begins by differentiating this sexual dysfunction from other sexual problems (e.g. pain causing interruption of intercourse), and by reviewing the conditions under which the patient is able to ejaculate (e.g. during sleep, with masturbation, with manual or oral stimulation). Generally, a complete evaluation should identify all possible predisposing, precipitating and maintaining factors for the dysfunction, such as issues of religiosity, coital and masturbatory patterns and/or performance anxiety.
Retrograde ejaculation
Often, a diagnosis of retrograde ejaculation can be made by taking an accurate history of previous surgical procedures and drug consumption. Retrograde ejaculation can be confirmed by demonstrating sperm presence in postcoital urine.8 A differential diagnosis should be pursued for patients who present with absent or low-volume ejaculates (such as anejaculation, obstruction of an ejaculatory duct or seminal vesicle, or congenital anomalies of the accessory sex organs), with a complete physical examination and transrectal ultrasonography.
Treatment of ejaculatory disorders
Premature ejaculation
The evolution of PE therapy has been influenced by new knowledge on physiology and etiology, and now includes behavioral, psychological and pharmacological therapies.
Currently, the pharmacological management of PE utilizes systemic or topical medications, either taken as needed or on a regular schedule. Apart from a novel agent, dapoxetine hydrochloride, the other remedies for PE are used in an off-label manner, as they were originally developed for the treatment of other pathologies but were secondarily found to delay ejaculation.
Among the various agents investigated for the oral treatment of PE, selective serotonin reuptake inhibitors (SSRIs) have demonstrated a variable efficacy compared to placebo.50 SSRIs are currently indicated as antidepressants, but citalopram, fluoxetine, fluvoxamine, paroxetine, and sertraline have also shown benefit in delaying ejaculation. In a meta-analysis of all treatment studies for PE, the majority of trials were found to lack an adequate design; only eight (18.5%) of the studies were performed according to the established evidence-based medicine criteria.59
The numerous trials studying the use of SSRIs for the management of PE have also evaluated daily versus on-demand dosing; however, the American Urological Association guidelines panel was not able to recommend a preferred dosing regimen and expressed a particular concern about the adverse-effect profiles of the various agents.50
According to results from placebo-controlled studies, daily treatment with sertraline, paroxetine, fluoxetine60 or clomipramine61 could yield safe benefits for men with PE. Above all, daily use of paroxetine has been demonstrated to yield the strongest ejaculatory delay within 5–10 days: the group with baseline IELT <1 href="http://www.nature.com/ncpuro/journal/v5/n2/full/ncpuro1016.html#B11">11 During treatment with SSRIs, adverse effects are usually mild to moderate and include fatigue, yawning and nausea.60 The loss of libido, ED, or significant agitation are rarely reported. The major contraindication for the daily use of SSRIs is the presence or a reported history of bipolar depression.
Waldinger et al.62 evaluated the preference of daily versus on-demand use of medication for PE in a group of 88 men with lifelong PE. Seventy one of the group (81%) preferred taking their medication daily; this result did not change after the patients were given the standard information about efficacy and adverse effects.62
On-demand administration of SSRIs 4–6 h before sexual intercourse is reported to be efficacious and well tolerated, but it is associated with a smaller delay in IELT than seen with daily administration. McMahon and Touma63 examined two placebo-controlled, crossover trials that investigated the effectiveness of paroxetine administered on a need-to-use basis. A significant IELT increase was registered after 3–4 weeks of use (P <0.001). href="http://www.nature.com/ncpuro/journal/v5/n2/full/ncpuro1016.html#B64">64 compared as-needed paroxetine to as-needed clomipramine with a stopwatch assessment in a randomized study. Paroxetine showed a 1.4-fold increase in IELT (95% CI 1.22–1.63), in contrast with a 4.0-fold increase when clomipramine was taken (95% CI 3.26–5.02); however, an elevated incidence of adverse effects, such as fatigue, sleepiness, dry mouth, tremor, yawning, loose stools, dizziness and headache, was reported after daily administration of both drugs.
Several rapidly acting and short half-life SSRIs are currently being studied for the treatment of PE. Oral dapoxetine hydrochloride is currently being considered for approval as the first pharmaceutical therapy developed with a specific indication for PE. In a 2007 double-blind, placebo-controlled trial, 212 potent men with PE were randomly assigned to receive either 30 mg oral dapoxetine twice daily or placebo.65 At the end of a 12-week treatment course, the dapoxetine group showed a 2.9-fold increase in mean IELT (95% CI 1.84–4.16). Significant increases were also observed in the mean weekly intercourse episodes and the mean intercourse satisfaction domain values, scored by the International Index of Erectile Function (IIEF) tool. Compared with placebo, dapoxetine use resulted in moderately prolonged IELTs and improved intercourse satisfaction, but lacked long-term benefits following discontinuation of the treatment. In a previous study, Pryor and colleagues66 assessed the efficacy of dapoxetine in two 12-week, randomized, double-blind, placebo-controlled, phase III trials of identical design. Men with moderate to severe PE received placebo, 30 mg dapoxetine or 60 mg dapoxetine as needed 1–3 h before sexual intercourse. Both dosages of dapoxetine significantly prolonged IELT compared with placebo (P <0.0001),>67 NO-mediated PDE-5 effects can thus target smooth muscle cells surrounding the vas deferens, seminal vesicles and prostate, improving the ejaculatory latency time.68
A multicenter, double-blind study was conducted with flexible doses (50–100 mg) of sildenafil in men with lifelong PE; a second group tested a single dose of 100 mg of sildenafil or placebo, taken 1 h before vibrotactile stimulation.67 Patients who received sildenafil reported increases in ejaculatory control and overall sexual satisfaction; however, the increased change in IELT did not reach statistical significance. Other studies investigated the use of combination therapies. Mattos and Lucon69 tested the use of tadalafil and the slow-release SSRI fluoxetine, alone or in combination therapy. Improved IELTs were obtained with the use of the agents in combination, compared with either single treatment. The efficacy of another PDE-5 inhibitor, vardenafil, was tested and compared with the SSRI sertraline; in men with primary or secondary PE, use of vardenafil resulted in longer IELTs than sertraline use.70
An alternative to oral medication is the use of topical agents to treat one of the supposed causes of PE—hypersensitivity of the penis. This approach involves applying local anesthetics in cream, gel or spray formulations in order to decrease penile sensitivity. Busato and colleagues71 designed a double-blind, randomized, placebo-controlled study to assess the use of lidocaine–prilocaine solution in 42 men with PE. A significant increase in mean IELT (P <0.001) href="http://www.nature.com/ncpuro/journal/v5/n2/full/ncpuro1016.html#B71">71, 72
Psychological treatment for PE has been proposed, with the aim of prolonging IELT and thus improving the sexual relationship between the patient and his partner. Several studies of psychotherapies have been reported, but there is a dearth of well-designed and controlled trials. Behavioral training techniques, including the stop-and-start and squeeze methods, are currently used by sexologists. The former was initially described by Masters and Johnson,9 and begins with manual autostimulation. After achievement of good arousal control by the man, the partner becomes involved with manual stimulation and then with sexual intercourse. The squeeze technique is similar to the aforementioned approach, except that when the stimulation is stopped, the penis is squeezed with the fingers.73 Although a good short-term success rate has been reported, behavioral techniques have never been studied in appropriate trials and tend to lose their efficacy over the long term.74
Some authors have tried to induce a permanent penile hypoesthesia by surgical selective dorsal nerve neurectomy. This surgical procedure has also been compared with hyaluronic acid gel injection, which can potentially block nerve receptors for tactile stimuli.75 No significant additive effect has been observed when injections and neurectomy are combined, but both individual treatments significantly prolong postoperative IELT.
Delayed ejaculation and anejaculation
Men suffering from delayed or inhibited ejaculation, when organic or pharmacologic causes are excluded, can benefit from a psychological approach. Preliminary psychosexual counseling is required before the psychotherapist can choose a therapy. Several approaches have been described: sex education, reduction of goal-oriented anxiety, increased and more genitally focused stimulation, and patient role-playing of an exaggerated ejaculatory response on his own and in front of his partner. In this field the literature is difficult to evaluate, as most authors report only a small series of cases with seemingly exaggerated results.76
A variety of drugs have been suggested to induce ejaculation, but no placebo-controlled studies have been published. Cyproheptadine, a serotonin-receptor antagonist, as well as several dopaminergic agents such as amantadine, yohimbine and apomorphine, are a few of the medications proposed but with variable and controversial results.
In order to achieve fertility for a patient who experiences anejaculation, sperm retrieval is possible via vibrostimulation.8 Vibrostimulation requires an intact lumbosacral spinal cord segment (above T10); if this method is not successful, electroejaculation, in which the periprostatic nerves are stimulated by a probe inserted into the rectum (generally under anesthesia), might be of benefit. If ejaculation does not occur using the aforementioned techniques, surgical sperm retrieval is the treatment of choice.
Retrograde ejaculation
When a spinal cord injury, urethral anomalies or drug consumption is excluded, men suffering from retrograde ejaculation can benefit from a pharmacological treatment.8 For example, imipramine, ephedrine sulfate and desipramine (chlorpheamine and phenylpropanalamine) have been found to improve bladder neck closure.77 Sperm collection from the urine is considered in cases of drug failure, contraindication for pharmacologic management, spinal cord injuries, or when other medications that induce retrograde ejaculation cannot be suspended.8
Conclusions
Neuropharmacological studies and animal research have contributed to an improved understanding of the neurobiology of ejaculation and the possible causes of ejaculatory disorders. Etiological treatments of underlying pathologies should be discussed with patients and offered as first-line therapies. SSRIs and topical anesthetic creams can provide good efficacy in treating PE, even if a clear understanding of the etiology of lifelong PE is lacking. Sperm retrieval techniques should be considered for patients with anejaculation or retrograde ejaculation who have associated infertility problems.
Key points
Ejaculatory dysfunction is one of the most common male sexual disorders, but is still frequently misdiagnosed
The most common ejaculatory dysfunction is premature ejaculation (PE), which affects 5–40% of sexually active men
Ejaculation involves cerebral sensory areas, motor centers and several spinal nuclei that are tightly interconnected
Diagnosis of PE in clinical practice is straightforward, as it is simply based on patient self report, clinical history, sexual history and examination findings
The major objective when diagnosing PE is to quantify the length of time between penetration and ejaculation, although a multidimensional assessment of patients affected with PE, including psychosocial involvement, is also needed
Selective serotonin reuptake inhibitors and topical anesthetic creams can provide good efficacy for treating PE, even if a clear understanding of the etiology of lifelong PE is lacking
References
Rosen R et al. (2003) Lower urinary tract symptoms and male sexual dysfunction: the multinational survey of the aging male (MSAM-7). Eur Urol 44: 637–649 Article PubMed ISI McMahon CG et al. (2006) Ejaculatory disorders. In Standard Practice in Sexual Medicine (Eds Porst H and Buvat J) Oxford: Blackwell Publishing
Jannini EA et al. (2002) Sexological approach to ejaculatory dysfunction. Int J Androl 25: 317–323 Article PubMed ISI ChemPort
Dunsmuir WD et al. (1996) There is a significant sexual dissatisfaction following TURP. Br J Urol 77: 161A
Rowland DL et al. (2001) Defining premature ejaculation for experimental and clinical investigations. Arch Sex Behav 30: 235–253 Article PubMed ISI ChemPort

ETICA DEL MEDICO COMO AGENTE FARMACEUTICO

DE LA VIDA MEDICALIZADA A UNA SOCIEDAD HIPOCONDRIACA

NEW ENG J MED.Volume 357:1796-1797
November 1, 2007
Number 18
Doctors and Drug Companies — Scrutinizing Influential Relationships
Eric G. Campbell, Ph.D.
On September 6, 2007, Senators Charles Grassley (R-IA), the ranking member of the Committee on Finance, and Herb Kohl (D-WI), chairman of the Special Committee on Aging, introduced the Physician Payments Sunshine Act — so named because it aims to "shine a much needed ray of sunlight on a situation that contributes to the exorbitant cost of health care," according to cosponsor Senator Charles Schumer (D-NY). The bill would require manufacturers of pharmaceuticals and medical devices with annual revenues of more than $100 million to disclose the amount of money they give to physicians — whether in the form of a free dinner or vacation or a consulting fee. "This bill is about letting the sun shine in so that the public can know," says Grassley.
The move was stimulated in part by activity in Minnesota and Vermont, which have made the reporting of such relationships mandatory — Minnesota in 1993 and Vermont in 2003. Three additional states (Maine, West Virginia, and California) and the District of Columbia have now enacted similar disclosure laws, and many other states are considering doing so. Although beliefs vary widely about the overall usefulness of the data collected under state mandates, the movement toward increased transparency is gaining steam.
Indeed, the nature, extent, and consequences of physicians' relationships with industry have become one of the most fiercely debated issues in health care today. At the simplest level, such a relationship exists whenever a physician accepts anything from a company whose products or services are related to the practice of medicine. And such interactions are ubiquitous: according to a recent survey, although the frequency and intensity of the ties vary according to physicians' personal and professional characteristics, virtually all physicians (94%) have some type of relationship with industry.1 Most commonly, physicians report receiving food and beverages in the workplace (83%) or being given drug samples by a manufacturer's representative (78%). More than one third of physicians (35%) receive reimbursement for costs associated with professional meetings or continuing medical education, and more than one quarter (28%) receive payments for consulting, speaking, or enrolling patients in trials 1
Frequency of Various Types of Physician–Industry Relationships.
Percentages were weighted to adjust for the probability of selection within each specialty and for nonresponse. Data are from Campbell et al.1
From a policy perspective, the debate centers on the overall effect of these relationships on patient care. Although most physicians deny that receiving free lunches, subsidized trips, or other gifts from pharmaceutical companies has any effect on their practices, research has shown that physician–industry relationships do influence prescribing behavior.2 After all, if these relationships didn't affect physician behavior in such a way as to increase sales, companies wouldn't spend $19 billion each year establishing and maintaining them.
Clearly, relationships between physicians and industry can have some positive effects on patient care. Many, if not all, of the drugs currently on the market simply wouldn't exist if it weren't for relationships whereby physicians enroll patients in manufacturers' clinical trials and provide companies with advice on drug development. And even a relationship in the form of attendance at an industry-sponsored luncheon or dinner may lead physicians to recommend beneficial drugs that are being underprescribed — though there is no reason why an educational activity needs to be accompanied by an expensive meal or a trip to a tropical resort.
But physician–industry relationships can also have serious negative effects. For example, doctors with ties to industry may be more inclined than their colleagues to prescribe a brand-name drug despite the availability of a cheaper generic version. The provision of free samples may reinforce this behavior and perhaps stimulate off-label use of medications, which can pose risks for some patients. Industry relationships may stimulate the premature adoption of novel treatments, which could lead to serious health problems for patients. Industry inducements may reduce physician adherence to evidence-based practice guidelines in favor of company medications or interventions that are not recommended in independently developed guidelines. Finally, the financial rewards from industry relationships may reinforce a culture of entitlement among physicians, which could limit their ability to honestly acknowledge and manage the potential negative effects of these relationships. In general, physicians vehemently deny that their industry relationships have any of these negative effects — but they are less convinced that the same is true of their physician colleagues.3
Concern about the risks associated with physician–industry relationships has inspired action by policymakers at the federal, state, and institutional levels. The same day that the new Senate bill was introduced, Boston University School of Medicine and Boston Medical Center announced a new conflict-of-interest policy — prohibiting their clinicians from accepting gifts from manufacturers of pharmaceuticals and medical devices, banishing industry-funded meals from campus, and requiring physicians who serve on the hospital's drug-selection committees to be free of financial ties to companies that stand to gain from committee decisions. Similar policy changes designed to limit physician–industry relationships have been implemented at institutions such as the University of Michigan Health System, the University of Pennsylvania, the Yale University School of Medicine, the Geisinger Health System in Pennsylvania, the Stanford University School of Medicine, Affinity Health System in Wisconsin, Kaiser Permanente, and HealthPartners in Minnesota. Such organizations have taken a variety of steps to banish drug representatives from their hospitals, reduce the use of drug samples accepted by physicians, eliminate industry-sponsored lunches, punish company representatives who break the rules (by loitering in a hospital, for instance, or giving out food), and educate doctors about pharmaceutical developments through academic physicians rather than industry representatives.
With other institutions now following suit, it seems likely that more stringent disclosure requirements and restrictions on industry relationships may soon become widespread — especially among physicians working in medical schools, teaching hospitals, and large health systems. Industry marketing practices such as sponsoring lunches at hospitals, offering free "educational" dinners at expensive restaurants, giving away trinkets such as coffee mugs and pens, and paying for questionable consultancy positions may well become much less common in the future.
Even as institutions and states exert increasing control over these relationships, such change has seemed unlikely to be implemented at the federal level, given the strength of the pharmaceutical-industry lobby in Washington. But in light of the Sunshine Act, federal regulation is clearly not impossible. The profession of medicine may view this legislation as a further stimulus to police itself more vigilantly, to work through its professional and specialty organizations to facilitate and monitor physician adherence to current policies regarding relationships with industry and, where necessary, to develop new, more stringent policies.
Individual physicians can take some steps to maximize the benefits for patients and minimize the risks associated with their own industry relationships. They can start by recognizing that such relationships are designed to influence prescribing behavior and by carefully considering the potential effects that their own associations may have on their patients. They can familiarize themselves with and adhere to the guidelines established by the institutions in which they practice and the professional associations to which they belong. And they can bear in mind that the costs of industry dinners, trips, and other incentives are passed along to their patients in the form of higher drug prices.
References
Campbell EG, Gruen RL,
.Mountford J, Miller LG, Cleary PD, Blumenthal D. A national survey of physician-industry relationships. N Engl J Med 200

LA JORNADA, 20 DE FEBRERO 2008
Arnoldo Kraus
Urdir enfermedades
Hay dos formas de inventar enfermedades. Una la hacen los individuos para satisfacer, modificar u ocultar alguna necesidad. La otra la generan los medios de publicación aliados e impulsados por las industrias farmacéuticas. La primera afecta, en general, sólo a una persona, es un reto médico y con frecuencia filosófico. La segunda perjudica a muchas personas y su génesis es doble: intereses económicos y vulnerabilidad médica. Escribo acerca de la segunda: la invención o exageración de enfermedades (en inglés, disease mongering) por parte de las compañías farmacéuticas es triste y cotidiana realidad.
Hace muchos años leí en una prestigiosa revista de medicina que lo peor que le podría pasar a la profesión médica era que los abogados se inmiscuyesen entre enfermos y doctores. Quien lo escribió tenía razón: el ejercicio médico, sobre todo en países como Estados Unidos, se ha desvirtuado y ha perdido su humanismo desde que los abogados tomaron algunas de las riendas de la profesión. Su presencia ha fracturado uno de los bienes más preciados de la profesión: la relación médico-paciente. Si la misma persona hubiese podido escribir en los últimos años un nuevo editorial diría que su profecía se cumplió, y agregaría que algo peor que la presencia de los abogados podría ser la irrupción de las compañías farmacéuticas y del poder de los medios de comunicación como enemigos de la profesión y de la comunidad.
Tanto abogados como medios de comunicación y farmacéuticas son constantes necesarias. El problema es el peso que han adquirido por culpa de la tibieza de los médicos y de los sistemas de salud, cuya fuerza y oposición a los dictados, no siempre deontológicos de las farmacéuticas, ha sido inconstante, mediocre y frágil.
La necesidad de urdir enfermedades o de medicarse se incrementa sin coto. Los medios de comunicación, maestros en disfrazar o inventar realidades, han dado en el clavo: cada vez es mayor la avidez de la sociedad para “conocer” y saber más acerca de su estado de salud. La retroactividad de la “arrogancia” de la medicina preventiva tal como la malentienden y la venden a los consumidores las compañías farmacéuticas en la televisión y en otros medios es nefanda: a mayor número de anuncios acerca de los milagros de las medicinas, mayor dependencia. La aspiración a la inmortalidad y a sufrir el menor número de enfermedades es leitmotiv de muchas personas, ese terreno ha sido explotado con maestría por la industria farmacéutica.
La invención o exageración de enfermedades con el propósito de mercantilizar productos farmacéuticos (disease mongering) implica crear padecimientos y ampliar los límites de las enfermedades con la finalidad de expandir los mercados y ofrecer tratamientos (he modificado la versión propuesta por Ray Moynihan y David Henry). Huelga decir que la invención de enfermedades es uno de los mayores logros de las grandes farmacéuticas, que, como se sabe, compiten en el mercado globalizado por la generación de dinero con el narcotráfico y con las empresas que fabrican armas.
La regla de oro para inventar o exagerar enfermedades es sencilla: transformar personas sanas en pacientes. Los dividendos son enormes: dinero a raudales por la dependencia que surge de la venta de medicamentos o del sentirse enfermo. Las consecuencias son terribles: se desperdician recursos y se genera iatrogenia (enfermedades producidas por los médicos). Y el final palpable: sobre todo en las sociedades ricas, cada vez hay más sanos transformados en enfermos que ingieren cotidianamente incontables químicos.
El problema es inmenso y seguramente irresoluble. Es inmenso, porque, aunque no cuento con “datos duros”, es evidente que el número de horas en la televisión o el número de anuncios en periódicos y en radio de las industrias farmacéuticas es muy grande y es irresoluble porque ningún medio informativo despreciará el negocio. Cada vez es más frecuente la automedicación y no es raro, sobre todo en la consulta privada que atiende a la clase económica privilegiada, encontrar enfermos sanos que ingieren “muchos” productos promovidos en la mass media.
Apelando a la filosofía, podría decirse que la apuesta de algunas compañías farmacéuticas es vender enfermedades en lugar de educar o de fomentar la prevención. En este entramado es imposible olvidar al finado Ivan Illich, quien hace más de tres décadas advertía que la profesión médica había medicalizado la vida. La suma de la medicalización de la vida por parte de la timorata profesión médica y la invención de enfermedades como estrategia de las industrias farmacéuticas es alarmante. Ejemplos sobran: la osteoporosis y los niveles de colesterol elevados se han convertido en enfermedades, la necesidad de recurrir a exámenes onerosos en ritual médico y la disfunción sexual masculina y femenina en negocio millonario.
La invención de enfermedades es otra de las enfermedades de la modernidad y triunfo de la comercialización de la vida sobre la ética que debería regir el mundo de la medicina. Dado que el espacio, no las denuncias, se ha agotado, expandiré estas diatribas la próxima semana.